CCHM2LAB W17 B.

Zimmerman Reaction → measures steroids with 17-keto structure,

uses M-dinitrobenzene to form reddish-purple complex
Hormone Assay C. Pisano Method → for quantitating metanephrines and
normetanephrines
D. Kober Method → for estrogen
SAMPLES FOR HORMONE ASSAY − H2SO4 + hydroquinone = reddish-brown color
Whole Blood – Ex. Luteinizing Hormone (LH), testosterone
Plasma LABORATORY MEASUREMENT:
→ EDTA – ACTH, ADH, PTH HORMONES OF THE THYROID GLAND
→ Heparin – Catecholamines, cortisol, dopamine, FSH
Serum – aldosterone, androstenedione, DHEA, estrogen, FSH, GH, HCG, Serum Free Triiodothyronine
progesterone » Measures the amount of free triiodothyronine (T 3) in blood
Urine – estriol » Measured using RIA or chemiluminometric assay
→ Boric Acid (1g/dL) – estriol and estrogen → 7 days » Used to evaluate and manage disorders of the thyroid gland
→ 10 mL of 6N HCl – 3-4 L container → catecholamines, VMA, 5- » Related tests: TSH, FT4
HIAA
→ HCl → maintains urine pH of <3.0 → chemical testing  Adults: 1.4-4.4 pg/mL (0.22-6.78 pmol/L)
 N>37 weeks (cord blood): 15-391 pg/dL (0.2-6 pmol/L)
I. CLASSIC ASSAYS  Pregnancy, 1st: 211-383 pg/dL (3.2-5.9 pmol/L)
A. Bioassays  Pregnancy, 2nd: 196-338 pg/dL (3-5.2 pmol/L)
− Based on observations of the physiological responses specific  Pregnancy, 3rd: 196-338 pg/dL (3-5.2 pmol/L)
for the hormone being measured
− Involves injection of test materials into prepared animal Direct Equilibrium Dialysis
− Ex. Galli mainini method → for pregnancy testing; uses male » Measures the amount of free thryoxine (T 4)
toad » Uses undiluted serum dialyzed for 16-18 hours at 37°C
» Dialysate is then analyzed directly using RIA
B. Competitive Protein Binding
− Based on competition for protein binding sites between tagged  2-128 ng/L (2.6 to 165 pmol/L)
or labeled hormone and the unlabeled hormone in the
patient’s sample Ultracentrifugation
− Ex.: Total T4 → Based on the specific binding properties of » How FT3 or FT4 are separated from the sample.
TBG (thyroxine-binding globulin) » Serum is adjusted first to pH of 7.4
» Incubated for 20 minutes at 37°C
II. IMMUNOLOGIC ASSAYS » ULTRACENTRIFUGE for 30 minutes at 37°C and 2000 rpm
➢ MOST WIDELY USED method for quantitating hormone in w/c a » Ultrafiltrate is analyzed by immunoassay
labeled antibody is used to determine w/ non-isotopic label. » Less time consuming than dialysis
A. RIA − CPB (competitive protein-binding)
(Radioimmunoassay) technique that utilizes radio-labeled Triiodothyronine Measurement
hormones as the tagged hormone » Measures the level of total T3 in blood
and anti-sera prepared against the » Used to evaluate and manage thyroid dysfunction, including
specific hormone as binding site. hyperthyroidism
» Related tests: FT4, T3 uptake
B. IRMA − Radiolabeled substance is attached
(Immunoradiometricassay) to the antibody instead of hormone;
 Adults: 60-181 ng/dL (0.92-2.78 nmol/L)
based on Ag-Ab reaction.
 Pregnancy (last 5 mos): 116-247 ng/dL (1.79-3.8 nmol/L)
C. ELISA − Enzyme is attached to the antibody;
(Enzyme-Linked can be measured
Serum Total T4 Competitive Immunoassay
Immunosorbentassay) spectrophotometrically
» Measures the total amount of thyroxine/T4 (BOTH free and CHON
D. EMIT − Enzyme tags are used; enzyme is bound) in blood
(Enzyme Multiplied attached to the hormone or drug » Uses barbital buffers (vs TBPA [thyroxine binding pre-albumin]) and
Immunosorbent being tested 8-anilino-1-naphthalene-sulfonic acid (vs TBG [thyroxine binding
Technique) ▪ Rate of NADH produced is globulin])
directly proportional to the
amount of drug tested.  Adults: 4.5-10.9 µg/dL (58-140 nmol/L)
▪ Requires NO separation of
bound and free antigen Anti-TSH Receptor Autoantibody
E. Immunometric − TSH test » For diagnosis of Grave’s disease
− Sensitive Test » Detects autoantibodies that interfere with the binding of TSH to TSH
▪ Immunochemiluminescent receptor
» Serum + TSH receptor + I125 labelled TSH tracer
» Amount of free tracer is measured
III. FLUORESCENT TECHNIQUES
A. FPIA (Fluorescence Polarization Immunoassay)  Lower than 9 U/L
− fluorescein-labeled drug, serum, and antibody are mixed and
placed in the light path of a fluorometer Thyrotropin Releasing Hormone (TRH) Stimulation Test
B. Antibody-bound conjugate (polarized fluorescence) is inversely » Specific test for thyroid disorder.
proportional to serum drug concentration. » Injection of TRH and measurement of the output of TSH
» Used in the diagnosis of combined pituitary- thyroid disorders
IV. HPLC (High-Performance Liquid Chromatography) » Differentiates 2° hypothyroidism and 3° hypothyroidism
➢ It is based on the differential partitioning of the compounds
between the mobile phase and the stationary phase. TEST FOR THYROID FUNCTION
➢ Separation of hormones from the other components of the blood. TRH Stimulation Test
= The MOST SPECIFIC and SENSITIVE TEST
V. COLORIMETRY = Measures relationship between TRH and TSH
➢ Chemical reactions producing color production are being used for = Helpful in the detection of thyroid hormone resistance syndromes.
detection of hormones.
 DOSE NEEDED: 500 ug TRH by IV
A. Porter-Silber Method → detect 17-OHCS (17-  INCREASED LEVELS: Primary hypothyroidism
Hydroxycorticosteroid)  DECREASED LEVELS: Hyperthyroidism
− Phenylhydrazine will be mixed w/ sulfuric acid and alcohol to
form a yellow end color
Other laboratory tests:
▪ RAI (Radioactive Iodine) uptake (RAIU): FREE THYROXINE INDEX (FT4I)
 Based on the ability of the thyroid to = INDIRECTLY assesses the level of free T4 in blood.
concentrate, convert and release I2 = Based on the equilibrium relationship of bound T4 and FT4
 Measure the ability of the thyroid = Elevated in hyperthyroidism and decreased in hypothyroidism.
gland to trap iodine.
 Helpful in establishing the cause of hyperthyroidism = FT4I = TT4 x T3U (%) OR TT 4 x THBR
 HIGH UPTAKE indicates metabolically active gland 100

▪ TBI: T3 UPTAKE
 Based on the thyroid hormone transport system indirectly = Measures the number of available binding sites of the thyroxine
measuring the amount of TBG binding proteins most notably TBG.
= DO NOT measure the level of thyroid hormone in serum, BUT reflects
level of TBG.
▪ PBI (Protein-Bound Iodine) test:
 Based on thyroid hormone concentration representing the organic  INCREASED LEVELS: Hyperthyroidism
fraction of blood iodine that precipitates with serum proteins  DECREASED LEVELS: Hypothyroidism

▪ BMR (Basal Metabolic Range):

 Based on metabolic response measuring the O2 consumption in the
resting fasting state Test and Methods for the detection tumors involving the thyroid
glands:

Thyroxine Binding Globulin (TBG) Fine Needle Aspiration → MOST ACCURATE TOOL in the evaluation of
= Used to confirm result of FT3 and FT4, and TT4 and THBR thyroid nodules
= TBG excess leads to increased T3 and T4 but the unbound form will o The routine use prompts the identification and treatment of
remain unchanged. malignancies involving the thyroid gland.
= Estrogen increases TBG; androgens depress TBG o Avoids unnecessary surgeries in those w/ benign cases.
o A small gauge needle will be inserted into a nodule and the cells will
 INCREASED: Euthyroidism, pregnancy and estrogen surge be further aspirated for histopathological examination.
 DECREASED: anabolic steroids and nephrosis
 REFERENCE VALUE: 13-39 ug/dL Recombinant Human TSH→ use to test patients with thyroid cancers for
the presence of residual or recurrent disease.
TSH Immunoassay
= Measures the amount of thyroid stimulating hormone (TSH) in blood Serum Calcitonin → TUMOR MARKER for detecting residual thyroid
= Using chemiluminescence w/ low detection limit = VERY SENSITIVE metastasis in medullary thyroid carcinoma
= Related tests: T3 and T4 o Measured both before and after 6 months of surgery.
Adults: 0.5-4.7 µunits/L
Pregnancy (1st): 0.3-4.5 µunits/L Pentagastrin Stimulation Test → for diagnosis of medullary thyroid
Pregnancy (2nd): 0.5-4.6 µL carcinoma
Pregnancy (3rd): 0.8-5.2 µL

▪ TSH Test SUMMARY OF THYROID DISORDERS AND LABORATORY TESTS

 The MOST IMPORTANT thyroid function test; BEST
SCREENING TEST
 Helps in the early detection of hypothyroidism
 Used to differentiate primary hypothyroidism from
secondary hypothyroidism
 Used to adjust and monitor thyroid hormone replacement
therapy.

 INCREASED: primary hypothyroidism and Hashimoto’s

thyroiditis and thyrotoxicosis
 ELEVATED: px with TSH antibodies & those w/ thyroid DISORDER T3 T4 TSH FT4 rT3 Tg TBG
hormones resistance Grave’s Disease ↑ ↑ ↓ ↑ ↑ ↑ N
 DECREASED: primary hyperthyroidism & secondary and Primary N/ ↓ ↓ ↑ ↓ ↓ N/ ↓ N
tertiary hypothyroidism Hypothyroidism
 BELOW THE NORMAL: had treatment for Grave’s disease Hashimoto N/ ↓ N/ ↓ ↑ N/ ↓ ↓ N/ ↓ N
and euthyroidism
Thyroiditis
THYROGLOBULIN ASSAY (TG ASSAY) Nonthyroidal illness ↓ N/ ↓ V V N/ ↑ N N
= Normally used as a postoperative marker of thyroid cancer Thyroid Hormone ↑ ↑ N/ ↑ ↑ ↑ ↑ N
= If used as tumor marker, it is important to test for thyroglobulin Resistance
antibodies Neonatal ↓ ↓ ↑ ↓ ↓ N/ ↓ N
= Used in monitoring course of metastatic or recurrence of thyroid
Hypothyroidism
cancer.

 INCREASED: Used for differentiating subacute thyroiditis

 DECREASED: Thyrotoxicosis Factitia N = normal
V = variable
= METHODS FOR TESTING:
Double antibody RIA, ELISA, IRMA, ICMA

REVERSE T3 (RT3)
= Used to assess borderline or conflicting laboratory results.
= Considered as the 3rd major circulating thyroid hormone.
= Formed by removal of one iodine from the inner ring of T4 and end
product of T4 metabolism
= Used for identifying patient w/ euthyroid sick syndrome in which the
RT3 will be ELEVATED.
 REFERENCE VALUES: 38 - 44ng/dL
CCHM2LAB W16 Allow measurement of both liquid and solid

Trace and Toxic Elements

samples; However, the volatility of the
compound to be measured will be dependent.
To overcome this, chemical modifiers such as
Palladium nitrate, Magnesium nitrate, or
SAMPLE COLLECTION AND PROCESSING mixture of both can be used.
• Specimens must be collected with scrupulous attention to details such as 2) Flame
anticoagulant, collection apparatus, and specimen type (Urine sample,
Whole blood sample, Serum sample)
• Elements are of low concentration in biologic specimens and ubiquitous in INDUCTIVELY COUPLED PLASMA MASS SPECTROMETRY (ICP-MS)
environment.
▪ A state-of-the-art analytic technique for elemental analysis
o Much care must be taken to avoid contaminating specimens.
o The term PLASMA pertains to an ionized gas such as Argon, where
• Special sampling and collection devices, specially cleaned glassware, certain portions of electrons will be rendered free
water and reagents of high purity must be used. ▪ Measures mass-to-charge ratio (m/z) of selected analyte ions
o Lab environment must be carefully controlled: placement of trace ▪ Includes ion source, m/z analyzer and ion detector
elements lab in separate room, use of sticky mats at doors, non- ▪ Argon plasma induced by ICP instruments generates high temperature
shedding ceiling tiles, controlled air flow, disposable booties (~6,000 to ~10,000 K) and serves several purposes:
✓ Dries droplets produced by nebulizer
INSTRUMENTATION AND METHODS ✓ Vaporizes dried particles
*Note: There is no single technique for testing of all trace/toxic elements, it varies
✓ Atomizes any molecular species
✓ Thermally ionizes atoms
ATOMIC EMISSION SPECTROSCOPY (AES) ▪ Quantitative analysis: best performed with the use of an Internal Standard
▪ Components of AE spectrophotometer: (usually an uncommon element such as Ytrium)
1. Source, sufficiently hot to produce excited-state species
(responsible for atomizing the compound to be measured in QUADRUPOLE MASS SPECTROMETER
order to produce excited-state species; the excited-state species
→ The typical mass spectrometer used for ICP-MS
will emit radiation upon relaxation back to its ground state)
→ The analyzer consists of four parallel conducting rods arranged in a square
2. Wavelength selecting device (monochromator) for spectral
array; applying the use of radio frequency (RF) and voltages to the rods
dispersion of radiation and separation of analytic line from other
→ Other ICP-MS instruments incorporate high-resolution mass
radiation (for dispersion and separation of the compound from
spectrometers.
others)
o These are usually “double focusing sector field” instruments.
3. Detector, permitting measurement of radiation intensity Such instruments separate ions of different m/z values via
▪ Liquid sample containing element is converted into an aerosol and
deflection in a magnetic field, with ions of greater m/z being
delivered into source, where it receives energy to emit radiation. deflected to a lesser degree than those of lower m/z.
▪ Intensity of emitted radiation is correlated to concentration of an analyte
and is basis for quantitation
▪ Simplified Schematic of AES: INTERFERENCES
SPECTROSCOPIC NON-SPECTROSCOPIC
+ Result from a spectral overlap − Matrix interferences; anything
with spectrum of target analyte interfering with atomization.
+ ICP-MS spectral interferences Properties of significance:
+ Polyatomic species whose m/z  Viscosity
may overlap m/z of target analyte  Ionized elements
(false elevations)  Carbon
+ Spectral interferences arise from Ways to overcome:
nearby elements in the periodic 1) Matrix matching
Source: Detector: table 2) Dilution of sample
1) Flame (up to 3,000 K) 1) Photographic film Strategies to deal with this:
− Fuel: Hydrogen or Acetylene 2) Photomultiplier tubes 1) Continuum source
− Oxidant gases: Air oxygen or 3) Array-based detector background corrector, or
Nitrous oxide system 2) Zeeman background
2) Plasma correction
+ Doubly charged ions
ATOMIC ABSORPTION SPECTROSCOPY (AAS)
▪ An analytic procedure for quantitation of elements through absorption of
optical radiation by free atoms in gas phase ELEMENTAL SPECIATION
▪ Spectra of atoms are line spectra, specific for absorbing elements. (Beer- ➢ Because toxicity of an element may vary by its chemical form, methods are
Lambert’s Law) needed to identify specific chemical forms
▪ Three most important components of AA spectrophotometer: ➢ HYPHENATED ANALYSIS: combination of complementary analytic
1. A source, in which the sample is atomized at a sufficient techniques to measure specific form of analyte
temperature to produce an excited-state species. Those species ➢ Liquid chromatography-ICP-MS (LC-ICPMS)
will emit radiation upon relaxation back to the ground state.
 Atomizer = where the atoms of element will be formed Arsenic: LC-ICP-MS, HG-GFAAS
2. A wavelength selecting device (monochromator), for the Copper: SEC-ICP MS
spectral dispersion of the radiation and separation of the Lead: GC-GFAAS
analytical line from other radiation. Selenium: SEC-GFAAS
3. A detector permitting measurement of radiation intensity. Zinc: AEC-ICP MS
▪ Simplified Schematic of AAS:
SEC = Size Exclusion Chromatography
GFAAS = Graphite Furnace AAS
AEC = Anion Exchange Chromatography

ALTERNATIVE ANALYTIC TECHNIQUES

❖ Neutron activation analysis, Voltametric methods (such as Anodic
Stripping Voltametry [ASV])
❖ Ion chromatography (For Copper, Iron, Zinc in serum and plasma
sample), Gas Chromatography–Mass Spectrometry (GC-MS) (for Cadium,
Light Source: Atomizer: Chromium, Cobalt, Lead, Selenium in urine, serum samples)
1) Hollow Cathode 1) Graphite tubes (commonly used) (flameless) ❖ Laser ablation ICPMS (LA-ICP-MS) (for solid samples)
Lamp (commonly → made up of high purity polycrystalline
used) electrographite and coated with pyrolytic
2) Electrodeless graphite (can be heated using electric
discharge Lamp current)
(recommended for Elements that use GFAAS:
volatile elements)  Selenium
 Cadmium
 Lead
CCHM2LAB W15 Liability of person who violates the CONFIDENTIALITY of records:

Legal Aspects of Drug

• IMPRISONMENT ranging from 6 months and 1 day to 6 years.
• Fine ranging from 1,000 to 6,000 pesos.
• Maximum penalty shall be imposed in addition to absolute perpetual
Testing disqualification from any public office when offender is a government official
or employee.

COMPREHENSIVE DANGEROUS DRUGS ACT OF 2002 (R.A. 9165) Sec 91. RA 9165
− Safeguard the state, its citizen from harmful effects of dangerous drugs. − Responsibility, liability of government officials/ employees in testifying as
− Enhance the efficiency of the law against dangerous drugs. prosecution witness in Dangerous Drug Cases:
− Enhance campaign against trafficking and use of dangerous drugs. “failure refusal to testify as witness hall be punished with IMPRISONMENT of not
− Re-integrate drug users back into the society. less than 12 years and 1 day to 20 years and fine of not less than 500,000 pesos
and administrative liability.”
Art III, Sec. 36: Authorized Drug Testing
− Drug testing shall be done by any government forensic lab or lab accredited REGULATORY FUNCTIONS
and monitored DOH. 1. Bureau of Health Facilities and Services (BHFS) also known now as
− DOH shall set the price/cost of drug testing. Health Facilities and Services Regulatory Bureau (HFSRB)
− Shall use two methods of testing (screening and confirmatory). 2. Department of Health (DOH)

MANDATORY DRUG TESTING Duties and Responsibilities of DOH (Sec 76, RA 9165)
Who are required to undergo MANDATORY drug testing? − License, accredit, establish and maintain drug testing network and labs.
a) Applicants for driver’s license (new & renewal). − Initiate conduct scientific research on drugs and drug control.
→ Repealed by RA 10586 2013, NO LONGER required to undergo − Encourage, assist and accredit private centers.
b) Applicants for firearm’s license. − Promulgate rules and regulations re DTL.
c) Applicants for permit to carry firearms outside of residence. − Set minimum standards for accreditation.
d) Officers and members of the military, police and other law enforcement − Oversee, monitor integration of rehabilitation centers.
agencies (annual drug testing). − Charge reasonable fees.
e) All persons who by nature of their profession carry firearms (e.g. security
guards). Under Sec. 36 of R.A 9165
f) Any person arrested for violations of the provisions of R.A. 9165, subjected “Authorized drug testing shall be done by any government forensic laboratories
to screening drug test within 24 hours. If positive, confirmatory testing or by any of the drug testing laboratories accredited and monitored by the DOH”
within 15 days after receipt of results (Sec. 38, R.A. 9165).
g) All candidates for public office whether appointed or elected both in Sec. 39 of R.A. 965:
“The DOH shall be the sole body, tasked to license and accredited drug testing
national or local government.
centers in each province and city.”
There shall be control regulations, licensing and accreditation division under
RANDOM DRUG TESTING
supervision of the DOH for this purpose.
Who may undergo random drug testing?
a) Students of Secondary and Tertiary schools (DECS/CHED to implement).
DRUG ABUSE AND PREVENTION PROGRAM (DAPP) ORGANIZATIONAL
REQUISITES:
CHART
✓ Pursuant to rules and regulations as contained in the school’s
student handbook.
✓ With notice to parents.
✓ Government shall bear the cost of drug testing (private or public
schools).
b) Officers and employees of public and private offices (domestic or
overseas).
Provided, required under the company’s work rules and regulations.
REQUISITES:
✓ Cost shall be borne by employer.
✓ Positive results shall be ground for suspension or termination,
subject to provisions of Art. 282 of the Labor code.

❑ All POSITIVE drug test results shall be subjected to confirmatory testing

using GC-MS or such modern, acceptable methods.
❑ And if found positive (after confirmatory testing)?
First offense Second offense
Minimum 6 months Imprisonment 6 years & 1 day
rehabilitation in a government to 12 years and fine of 50,000
center to 200,000 pesos (Sec. 15)

TEST RESULT
• Valid for one (1) year from date of issue [Sec. 36].
• May be used for other purposes [Sec. 36].
• POSITIVE screening lab test must be confirmed for it to be valid in court
[Sec. 38].
• Shall be signed by the ANALYST: HEAD OF THE LAB shall sign ALL test
results to be submitted to any government agency.
• For (+) results: original shall be given to client immediately; copies shall be
given to BHFS and the requesting agency; DTL shall retain copy → 3
copies
• Shall be in the form prescribed by BHFS.
ORGANIZATIONS RESPONSIBILITIES
• Confidential; ONLY GOVERNMENT AGENCIES shall be provided with a
DDB: Dangerous  Policy making body related to dangerous drugs.
copy upon WRITTEN REQUEST. Drugs Board  Composed of Undersecretary representatives
from different government departments.
Issuance of FALSE or FRAUDULENT Drug Test Result Undersecretary  Program manager for DAPP.
for Special  Concerned with all activities related to drug
“xxx who issues false or fraudulent drug test results knowingly, willfully or abuse (drug testing rehabilitation, prevention)
Concerns
through negligence shall suffer the penalty of IMPRISONMENT of 6 years and  Responsible for setting the directions
1 day up to 12 years and a FINE from 100,00 to 50,000 pesos.” formulation of plans, and implementation of
− Plus, REVOCATION OF LICENSE to practice the profession and program.
CLOSURE of drug testing center [Sec. 37].
NCFHD: National  Tasked to develop, encourage and establish DRUG TESTING LABORATORY (DTL)
Center for Health drug testing and rehabilitation centers in
Facilities DOH retained hospitals and facilities.
Development  Monitor its activities.
BHFS: Bureau of  Accredit, license and monitor drug testing and
Health Facilities rehabilitation centers.
Services  Formulate licensing and accreditation standards
for DTL and rehabilitation centers.
 Has mandate to close substandard DT and
Rehabilitation facilities.
BFAD: Bureau of  Approves the registration of drug test kits and
Food and Drugs reagents for marketing purposes.
 Mandate to recall substandard products and
cancel registration certificates of manufactures.
BHDT: Bureau of  Approve the registration of health devices
Health Devices (equipment, machines) used for health
and Technology purposes.
 License and accredit radiologic facilities.
CHD: Center for  Formerly the Regional Health Center. Screening DTL: Personnel Requirements
Health  Deputized by BHFS to: Head of laboratory (HOL)
Development ▪ Accept application for re-accreditation. Analyst
▪ Inspect DTL for accreditation purposes. Authorized specimen collector (ASC)
▪ Monitor compliance to standards.
 Deputized by BFAD to: HEAD OF THE LABORATORY - Qualifications:
▪ Withdraw drug test kits from DTL for post
→ Screening laboratory:
market surveillance (future plans).
 Clinical pathologist
 Deputized by IMS to:
 Physician trained in laboratory management and quality assurance
▪ Register and validate drug kits for IDTOMIS
use. program
IMS: Information  Provides technical support in the IT → Confirmatory laboratory:
Management requirement of DTL and DRC.  Clinical pathologist
System  Formulates IT standards for DTL and DRC.  Chemist with knowledge and skills in operation of GC-MS
 Program implementer of DTOMIS and
IDTOMIS. ANALYST - Qualification
NRL: National  For environmental and occupational health, → Must be a registered:
Reference toxicology and micronutrient assay  Chemist
Laboratory  FUNCTIONS:  Medical technologist
▪ To provide laboratory referral services-  Pharmacist
confirmatory testing, surveillance &  Chemical engineer
research
- Confirmatory tests for drug abuse
AUTHORIZED SPECIMEN COLLECTOR
- Perform challenge test
▪ Develop & validate protocols of analysis for → A trained individual who instructs, assists a donor at collection site,
drugs of abuse. → receives and makes an initial inspection of specimen for drug testing
▪ To train laboratory personnel → initiates and completes the CCF.
- Develop training modules for technical → Qualifications:
personnel of DTL.  At least high school graduate
▪ Conduct training of analysts, HDL.  Must have undergone training on specimen collection
▪ To maintain a QAP for lab, tests in
coordination with the BHFS-DOH.
- Conduct of Proficiency tests (EQAS).
- Formulate standards of GLP in Drug
testing.
▪ To evaluate test kits & reagents, in
coordination with BFAD & the BHDT-DOH
and perform post market surveillance.
- Formulate technical protocol for drug
test kit validation.
- Perform technical evaluation of DKTR.
 During testing, a urine specimen migrates upward by capillary action. A drug, if present in the urine specimen INVALID: Control line fails to appear. Insufficient specimen volume or incorrect procedural techniques are
below its cut-off concentration, will not saturate the binding sites of its specific antibody. The antibody will the most likely reasons for control line failure. Review the procedure and repeat the test using a new test. If
Effective date: 2014-07-03 Number: 1156098001 then react with the drug-protein conjugate and a visible colored line will show up in the test line region. The the problem persists, discontinue using the lot immediately and contact your local distributor.
presence of drug above the cut-off concentration will saturate all the binding sites of the antibody. Therefore,
the colored line will not form in the test line region. QUALITY CONTROL
A drug-positive urine specimen will not generate a colored line in the test line region because of drug A procedural control is included in the test. A colored line appearing in the control line region (C) is
competition, while a drug-negative urine specimen will generate a line in the test line region because of the considered an internal procedural control. It confirms sufficient specimen volume, adequate membrane
One Step Drug Screen Test Device wicking and correct procedural technique.
absence of drug competition. To serve as a procedural control, a colored line will always appear at the control
(Urine) line region, indicating that proper volume of specimen has been added and membrane wicking has occurred. Control standards are not supplied with this kit. However, it is recommended that positive and negative
controls be tested as good laboratory practice to confirm the test procedure and to verify proper test performance.
Package Insert REAGENTS
Each test contains specific drug antibody-coupled particles and corresponding drug-protein conjugates. A LIMITATIONS
English goat antibody is employed in the control line. 1. The SPINREACT One Step Drug Screen Test Device (Urine) provides only a qualitative, preliminary
PRECAUTIONS analytical result. A secondary analytical method must be used to obtain a confirmed result. Gas
chromatography/mass spectrometry (GC/MS) is the preferred confirmatory method.2, 3
Package insert for testing of the following drugs: • For medical and other professional in vitro diagnostic use only. Do not use after the expiration date. 2. There is a possibility that technical or procedural errors, as well as other interfering substances in the urine
Amphetamine 300, Amphetamine 500, Amphetamine, Barbiturates, Benzodiazepines 200, Benzodiazepines, • The test device should remain in the sealed pouch until use. specimen may cause erroneous results.
Buprenorphine, Cocaine 150, Cocaine, Cotinine, Fentanyl, Ketamine, Marijuana 20, Marijuana, Marijuana • All specimens should be considered potentially hazardous and handled in the same manner as an 3. Adulterants, such as bleach and/or alum, in urine specimens may produce erroneous results regardless of the
150, Methadone, EDDP 100 (Methadone metabolite), EDDP 300 (Methadone metabolite), Methamphetamine infectious agent. analytical method used. If adulteration is suspected, the test should be repeated with another urine specimen.
300, Methamphetamine 500, Methamphetamine, Methylenedioxymethamphetamine, Morphine 300, Opiate • The used test device should be discarded according to local regulations. 4. A positive result does not indicate level or intoxication, administration route or concentration in urine.
2000, Oxycodone, Phencyclidine, Propoxyphene, Tramadol and Tricyclic Antidepressants. STORAGE AND STABILITY 5. A negative result may not necessarily indicate drug-free urine. Negative results can be obtained when drug
A rapid, one step screening test for the simultaneous, qualitative detection of drugs and drug metabolites in Store as packaged in the sealed pouch either at room temperature or refrigerated (2-30°C). The test device is is present but below the cut-off level of the test.
human urine. stable through the expiration date printed on the sealed pouch. The test device must remain in the sealed 6. The test does not distinguish between drugs of abuse and certain medications.
For medical and other professional in vitro diagnostic use only. pouch until use. DO NOT FREEZE. Do not use beyond the expiration date. 7. A positive result may be obtained from certain foods or food supplements.
INTENDED USE & SUMMARY SPECIMEN COLLECTION AND PREPARATION PERFORMANCE CHARACTERISTICS
Urine based tests for drugs of abuse range from simple immunoassay tests to complex analytical procedures. Accuracy
Urine Assay
The speed and sensitivity of immunoassays have made them the most widely accepted method to screen urine A side-by-side comparison was conducted using the SPINREACT One Step Drug Screen Test Device (Urine)
for drugs of abuse. The urine specimen must be collected in a clean and dry container. Urine collected at any time of the day may
and a commercially available drug rapid test. Testing was performed on approximately 300 specimens
The SPINREACT One Step Drug Screen Test Device (Urine) is a lateral flow chromatographic immunoassay be used. Urine specimens exhibiting visible precipitates should be centrifuged, filtered, or allowed to settle to
previously collected from subjects presenting for Drug Screen Testing. Presumptive positive results were
for the qualitative detection of drugs and drug metabolites in urine at the following cut-off concentrations in obtain a clear supernatant for testing.
confirmed by GC/MS. Negative urine specimens were screened initially by Predicate test, 10% negative
urine:1 Specimen Storage
specimens were confirmed by GC/MS. The following results were tabulated:
Cut-off (ng/mL) Urine specimens may be stored at 2-8°C for up to 48 hours prior to testing. For prolonged storage, specimens
Test Calibrator may be frozen and stored below -20°C. Frozen specimens should be thawed and mixed well before testing. % Agreement with Commercial Kit
Amphetamine (AMP 300) d-Amphetamine 300 AMP AMP BZO COC THC THC
MATERIALS Specimen
300 500
AMP BAR
200
BZO BUP**
150
COC COT FTY KET
20
THC
150
Amphetamine (AMP 500) d-Amphetamine 500
Amphetamine (AMP) d-Amphetamine 1,000
Materials Provided Positive >99% * 96% >99% * 90% 88% >99% 95% >99% * * * >99% *
Barbiturates (BAR) Secobarbital 300 • Test devices • Droppers • Package insert Negative >99% * >99% 99% * 97% >99% >99% >99% >99% * * * >99% *
Benzodiazepines (BZO 200) Oxazepam 200 Materials Required But Not Provided Total >99% * 98% 99% * 94% 97% >99% 98% >99% * * * >99% *
Benzodiazepines (BZO) Oxazepam 300 • Specimen collection container • Timer EDDP EDDP MET MET MOP OPI
Buprenorphine (BUP) Buprenorphine 10 Specimen MTD MET MDMA OXY PCP PPX TRA TCA
DIRECTIONS FOR USE 100 300 300 500 300 2000
Cocaine (COC 150) Benzoylecgonine 150 Positive >99% * * * >99% 99% >99% >99% 99% 96% 97% >99% * 95%
Allow the test, urine specimen, and/or controls to reach room temperature (15-30ºC) prior to testing.
Cocaine (COC) Benzoylecgonine 300 1. Bring the pouch to room temperature before opening it. Remove the test device from the sealed pouch and Negative >99% * * * 82% >99% 99% >99% >99% 99% >99% >99% * >99%
Cotinine (COT) Cotinine 100 use it as soon as possible. Total >99% * * * 89% >99% 99% >99% >99% 98% 99% >99% * 99%
Fentanyl (FTY) Norfentanyl 20 2. Place the test device on a clean and level surface. Hold the dropper vertically and transfer 3 full drops of * NOTE: Commercial kit unavailable for comparison testing.
Ketamine (KET) Ketamine 1,000 urine (approx. 100 μL) to the specimen well (S) of the test device, and then start the timer. Avoid trapping ** NOTE: BUP was compared to the self-reported use of Buprenorphine
Marijuana (THC 20) 11-nor-Δ9-THC-9 COOH 20 air bubbles in the specimen well (S). See the illustration below. % Agreement with GC/MS
Marijuana (THC) 11-nor-Δ9-THC-9 COOH 50 3. Wait for the colored line(s) to appear. Read results at 5 minutes. Do not interpret the result after AMP AMP BZO COC THC THC
Marijuana (THC 150) 11-nor-Δ9-THC-9 COOH 150 10 minutes. Specimen AMP BAR BZO BUP* COC COT* FTY* KET THC
300 500 200 150 20 150
Methadone (MTD) Methadone 300 Positive >99% 97% 96% 92% 98% 96% 98% 99% 96% >99% 99% >99% 91% 97% 91%
Methadone metabolite (EDDP 100) 2-Ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) 100 Negative 99% 99% 95% 98% 99% 96% >99% 99% 90% >99% 89% 97% 99% 96% 96%
Methadone metabolite (EDDP 300) 2-Ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) 300 Total 99% 98% 95% 95% 99% 96% >99% 99% 93% >99% 93% 97% 96% 97% 95%
Methamphetamine (MET 300) d-Methamphetamine 300
Methamphetamine (MET 500) d-Methamphetamine 500 EDDP EDDP MET MET MOP OPI
Specimen MTD MET MDMA OXY PCP PPX TRA* TCA**
100 300 300 500 300 2000
Methamphetamine (MET) d-Methamphetamine 1,000
Positive 99% >99% >99% 98% 99% 99% >99% >99% 98% 99% >99% 94% 96% >99%
Methylenedioxymethamphetamine (MDMA) d,l-Methylenedioxymethamphetamine 500
Negative 94% >99% 95% >99% 98% 93% 98% 94% 97% 98% 97% 99% 97% 89%
Morphine (MOP 300) Morphine 300
Total 96% >99% 97% 99% 98% 96% 99% 97% 98% 99% 98% 96% 97% 91%
Opiate (OPI 2000) Morphine 2,000
Oxycodone (OXY) Oxycodone 100 * NOTE: BUP, COT, FTY and TRA were based on LC/MS data instead of GC/MS.
** NOTE: TCA was based on HPLC data instead of GC/MS.
Phencyclidine (PCP) Phencyclidine 25
Propoxyphene (PPX) Propoxyphene 300 Analytical Sensitivity
Tramadol (TRA) Tramadol 100 A drug-free urine pool was spiked with drugs to the concentrations at ± 50% cut-off and ± 25% cut-off. The
Tricyclic Antidepressants (TCA) Nortriptyline 1,000 results are summarized below.
INTERPRETATION OF RESULTS
This test will detect other related compounds, please refer to the Analytical Specificity table in this package insert. Drug Conc. AMP 300 AMP 500 AMP BAR BZO 200 BZO BUP COC 150 COC COT
This assay provides only a preliminary analytical test result. A more specific alternate chemical (Please refer to the illustration above)
(Cut-off range) - + - + - + - + - + - + - + - + - + - +
method must be used in order to obtain a confirmed analytical result. Gas chromatography/mass NEGATIVE:* Two lines appear. One colored line should be in the control line region (C), and another
spectrometry (GC/MS) is the preferred confirmatory method. Clinical consideration and professional apparent colored line should be in the test line region (T). This negative result indicates that the drug 0% Cut-off 30 0 90 0 30 0 30 0 60 0 30 0 90 0 30 0 30 0 90 0
judgment should be applied to any drug of abuse test result, particularly when preliminary positive concentration is below the detectable level. -50% Cut-off 30 0 90 0 30 0 30 0 60 0 30 0 90 0 30 0 30 0 90 0
results are used. *NOTE: The shade of color in the test line region (T) will vary, but it should always be considered as -25% Cut-off 25 5 88 2 23 7 20 10 60 0 26 4 78 12 27 3 30 0 90 0
PRINCIPLE negative whenever there is even a faint colored line. Cut-off 16 14 45 45 9 21 13 17 22 38 12 18 48 42 13 17 9 21 49 41
The SPINREACT One Step Drug Screen Test Device (Urine) is an immunoassay based on the principle of POSITIVE: One colored line appears in the control line region (C). No line appears in the test line region +25% Cut-off 4 26 1 89 1 29 8 22 2 58 3 27 24 66 7 23 7 23 4 86
competitive binding. Drugs which may be present in the urine specimen compete against their respective drug (T). This positive result indicates that the drug concentration exceeds the detectable level. +50% Cut-off 0 30 0 90 0 30 0 30 0 60 0 30 0 90 0 30 0 30 0 90
conjugate for binding sites on their specific antibody.
English 1
 THC EDDP EDDP Desalkylflurazepam 1,560 p-Hydroxymethamphetamine 30,000 Methadone 300 Maprotiline 2,000
Drug Conc. FTY KET THC 20 THC MTD MET 300 MET 500
150 100 300 Diazepam 97 Mephentermine 50,000 Doxylamine 50,000 Nordoxepin 1,000
(Cut-off range)
- + - + - + - + - + - + - + - + - + - + Estazolam 125 l-Methamphetamine 8,000 MARIJUANA 20 Promazine 1,500
0% Cut-off 90 0 90 0 30 0 30 0 90 0 30 0 90 0 90 0 30 0 30 0 Flunitrazepam 25,000 d,l-3,4-Methylenedioxymethamphetamine (MDMA) 2,000 11-nor-Δ9-THC-9 COOH 20 Promethazine 25,000
-50% Cut-off 90 0 90 0 30 0 30 0 90 0 30 0 90 0 90 0 30 0 30 0 α-Hydroxyalprazolam 30 METHYLENEDIOXYMETHAMPHETAMINE (MDMA) Cannabinol 12,500 Trimipramine 3,000
-25% Cut-off 79 11 48 42 29 1 30 0 90 0 26 4 80 10 79 11 27 3 27 3 d-Lorazepam 3,125 d,l-3,4-Methylenedioxymethamphetamine (MDMA) 500 11-nor-Δ8-THC-9 COOH 20
Cut-off 36 54 6 84 19 11 21 9 45 45 16 14 51 39 51 39 15 15 13 17 Midazolam 195 d,l-3,4-Methylenedioxyamphetamine (MDA) 3,000 Δ8-THC 10,000
+25% Cut-off 7 83 0 90 6 24 17 13 10 80 4 26 3 87 13 77 5 25 7 23 Nitrazepam 780 3,4-Methylenedioxyethylamphetamine (MDEA) 300 Δ9-THC 12,500
+50% Cut-off 0 90 0 90 0 30 0 30 0 90 0 30 0 90 0 90 0 30 0 30 Norchlordiazepoxide 780 MORPHINE 300
Cross-Reactivity
Nordiazepam 780 Morphine 300
Drug Conc. A study was conducted to determine the cross-reactivity of the test with compounds in either drug-free urine or
MET MDMA MOP 300 OPI 2000 OXY PCP PPX TRA TCA Temazepam 33 Codeine 300
(Cut-off range) Amphetamine 300, Amphetamine 500, Amphetamine, Barbiturates, Benzodiazepines 200, Benzodiazepines,
Triazolam 150 Ethylmorphine 6,250
- + - + - + - + - + - + - + - + - + Buprenorphine, Cocaine 150, Cocaine, Cotinine, Fentanyl, Ketamine, Marijuana 20, Marijuana, Marijuana 150,
BENZODIAZEPINES Hydrocodone 50,000
0% Cut-off 30 0 30 0 30 0 30 0 30 0 30 0 30 0 90 0 30 0 Methadone, EDDP 100, EDDP 300, Methamphetamine 300, Methamphetamine 500, Methamphetamine,
Oxazepam 300 Hydromorphone 3,125 Methylenedioxymethamphetamine, Morphine 300, Opiate 2000, Oxycodone, Phencyclidine, Propoxyphene,
-50% Cut-off 30 0 30 0 30 0 30 0 30 0 30 0 30 0 90 0 30 0
Alprazolam 196 Levorphanol 1,500 Tramadol and Tricyclic Antidepressants positive urine. The following compounds show no cross-reactivity when
-25% Cut-off 24 6 23 7 28 2 24 6 30 0 26 4 26 4 90 0 26 4
Cut-off 18 12 15 15 20 10 10 20 21 9 11 19 19 11 58 32 14 16
Bromazepam 1,562 6-Monoacetylmorphine (6-MAM) 400 tested with the SPINREACT One Step Drug Screen Test Device (Urine) at a concentration of 100 μg/mL.
Chlordiazepoxide 1,562 Morphine 3-β-D-glucuronide 1,000 Non Cross-Reacting Compounds
+25% Cut-off 1 29 6 24 3 27 4 26 6 24 8 22 8 22 22 68 4 26
Clobazam 98 Norcodeine 6,250 4-Acetamidophenol Diclofenac Labetalol Prednisolone
+50% Cut-off 0 30 0 30 0 30 0 30 0 30 0 30 0 30 2 88 0 30
Clonazepam 781 Normorphine 100,000 Acetone Dicyclomine Lidocaine Prednisone
Analytical Specificity Clorazepate 195 Oxycodone 30,000 Acetophenetidin Diflunisal Lindane d,l-Propanolol
Delorazepam 1,562 Oxymorphone 100,000 Acetylsalicylic acid Digoxin Lithium Quinacrine
The following table lists the concentration of compounds (ng/mL) that are detected positive in urine by the
Desalkylflurazepam 390 Procaine 15,000 Albumin 4-Dimethylaminoantipyrine Loperamide Quinidine
SPINREACT One Step Drug Screen Test Device (Urine) at 5 minutes. alpha-Naphthaleneacetic Acid Diphenhydramine l-Thyroxine Quinine
Diazepam 195 Thebaine 6,250
AMPHETAMINE 300 MARIJUANA Aminopyrine 5,5-Diphenylhydantoin Meperidine R(-) Deprenyl
Estazolam 2,500 OPIATE 2000 Amoxapine EMDP Meprobamate Riboflavin
d-Amphetamine 300 11-nor-Δ9-THC-9 COOH 50
Flunitrazepam 390 Morphine 2,000 Amoxicillin Erythromycin Methaqualone Salicylic acid
d,l-Amphetamine 390 Cannabinol 20,000
α-Hydroxyalprazolam 1,262 Codeine 2,000 Ampicillin β-Estradiol Methoxyphenamine Serotonin
l-Amphetamine 50,000 11-nor-Δ8-THC-9 COOH 30 Apomorphine Estrone-3-sulfate Methylphenidate Seroquel
d,l-Lorazepam 1,562 Ethylmorphine 5,000
p-Hydroxyamphetamine 1,560 Δ8-THC 15,000 Ascorbic acid Ethyl alcohol Metoprolol Sertraline
RS-Lorazepam glucuronide 156 Hydrocodone 12,500
p-Hydroxynorephedrine 100,000 Δ9-THC 15,000 Aspartame Ethyl-p-aminobenzoate N-Acetylprocainamide Sodium Chloride
Midazolam 12,500 Hydromorphone 5,000 Atropine Etodolac Nalidixic acid Sulfamethazine
3,4-Methylenedioxyamphetamine (MDA) 1,560 MARIJUANA 150
Nitrazepam 98 Levorphanol 75,000 Benzilic acid Famprofazone Nalorphine Sulindac
β-Phenylethylamine 100,000 11-nor-Δ9-THC-9 COOH 150
Norchlordiazepoxide 195 6-Monoacetylmorphine (6-MAM) 5,000 Benzoic acid Fenoprofen Naproxen Tetracycline
Phenylpropanolamine (d,l-Norephedrine) 100,000 Cannabinol 25,000 Benzydamine Fluoxetine Niacinamide Tetrahydrocortison-3-acetate
Nordiazepam 390 Morphine 3-β-D-glucuronide 2,000
Tyramine 100,000 11-nor-Δ8-THC-9 COOH 500 Brompheniramine Furosemide Nifedipine Tetrahydrozoline
Temazepam 98 Norcodeine 12,500
AMPHETAMINE 500 Δ8-THC 25,000 Caffeine Gentisic acid Nimesulide Theophylline
Triazolam 2,500 Normorphine 50,000 Cannabidiol d-Glucose Norethindrone Thiamine
d-Amphetamine 500 Δ9-THC 25,000
BUPRENORPHINE Oxycodone 25,000 Chloral Hydrate Guaiacol Glyceryl Ether Noscapine Thioridazine
d,l-Amphetamine 1,500 EDDP 100
Buprenorphine 10 Oxymorphone 25,000 Chloramphenicol Hemoglobin d,l-Octopamine Tolbutamide
Methamphetamine 780 2-Ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) 100 Chloroquine Hydralazine Orphenadrine Trans-2-phenylcyclopropylamine
Buprenorphine 3-D-glucuronide 15 Procaine 150,000
p-Hydroxybuprenorphine 1,562 EDDP 300 Chlorothiazide Hydrochlorothiazide Oxalic acid Trazodone
Norbuprenorphine 20 Thebaine 100,000
l-Amphetamine 25,000 2-Ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) 300 Chlorpromazine Hydrocortisone Oxolinic acid Triamterene
Norbuprenorphine 3-D-glucuronide 200 OXYCODONE Chlorprothixene o-Hydroxyhippuric acid Oxymetazoline Trifluoperazine
AMPHETAMINE METHAMPHETAMINE 300
COCAINE 150 Oxycodone 100 Cholesterol 3-Hydroxytyramine Papaverine Trimethoprim
d-Amphetamine 1,000 d-Methamphetamine 300
Benzoylecgonine 150 Hydrocodone 6,250 Cimetidine Ibuprofen Pemoline d,l-Tryptophan
d,l-Amphetamine 3,000 d,l-Amphetamine 100,000
Cocaine 400 Hydromorphone 50,000 Clonidine Iproniazid Penicillin d,l-Tyrosine
l-Amphetamine 50,000 Chloroquine 25,000 Cortisone Isoproterenol Pentazocine Uric acid
Cocaethylene 6,250 Levorphanol 50,000
d,l-3,4-Methylenedioxyamphetamine (MDA) 2,000 Ephedrine 100,000 Creatinine Isoxsuprine Phenelzine Verapamil
Ecgonine 12,500 Naloxone 37,500
Phentermine 3,000 (1R,2S)-l-Ephedrine 100,000 Deoxycorticosterone Kanamycin Pheniramine Zomepirac
Ecgonine methylester 50,000 Naltrexone 37,500 Dextromethorphan Ketoprofen Phenothiazine
BARBITURATES l-Epinephrine 50,000
COCAINE Oxymorphone 200
Secobarbital 300 Fenfluramine 12,500 BIBLIOGRAPHY
Benzoylecgonine 300 PHENCYCLIDINE
Alphenal 150 p-Hydroxymethamphetamine 25,000 1. Tietz NW. Textbook of Clinical Chemistry. W.B. Saunders Company. 1986; 1735
Cocaine 780 Phencyclidine 25
Amobarbital 300 Mephentermine 50,000 2. Baselt RC. Disposition of Toxic Drugs and Chemicals in Man. 2nd Ed. Biomedical Publ., Davis, CA.
Cocaethylene 12,500 4-Hydroxyphencyclidine 12,500
Aprobarbital 200 l-Methamphetamine 3,125 1982; 488
Ecgonine 32,000 PROPOXYPHENE
Butabarbital 75 3,4-Methylenedioxymethamphetamine (MDMA) 780 3. Hawks RL, CN Chiang. Urine Testing for Drugs of Abuse. National Institute for Drug Abuse (NIDA),
COTININE d-Propoxyphene 300
Butalbital 2,500 Trimethobenzamide 25,000 Research Monograph 73, 1986
l-Cotinine 100 d-Norpropoxyphene 300
Butethal 100 METHAMPHETAMINE 500 Index of Symbols
S-l-Nicotine 12,500 TRAMADOL
Cyclopentobarbital 600 d-Methamphetamine 500
FENTANYL n-Desmethyl-cis-tramadol 195 Consult instructions
Pentobarbital 300 d,l-Amphetamine 75,000 Tests per kit Manufacturer
Norfentanyl 20 o-Desmethyl-cis-tramadol 6,250 for use
Phenobarbital 100 d-Amphetamine 50,000
Alfentanyl 562,500 Cis-tramadol 100 For in vitro
BENZODIAZEPINES 200 Chloroquine 12,500 Use by Do not reuse
Buspirone 12,500 Phencyclidine 100,000 diagnostic use only
Oxazepam 200 (1R,2S)-l-Ephedrine 50,000
Fenfluramine 37,500 Procyclidine 100,000
Alprazolam 30 p-Hydroxymethamphetamine 15,000 Store between 2-30°C Lot Number REF Catalog #
Fentanyl 100 d,l-O-Desmethyl venlafaxine 25,000
7-Aminoclonazepam 4,000 Mephentermine 25,000
Sufentanyl 57,500 TRICYCLIC ANTIDEPRESSANTS
7-Aminoflunitrazepam 390 l-Methamphetamine 4,000
KETAMINE Nortriptyline 1,000
7-Aminonitrazepam 625 3,4-Methylenedioxymethamphetamine (MDMA) 1,000 SPINREACT, SAU
Ketamine 1,000 Amitriptyline 1,500
Bromazepam 390 l-Phenylephrine 100,000 Ctra. Santa Coloma, 7, 17176 Sant Esteve de Bas, GIRONA. Spain
Pentobarbital 50,000 Clomipramine 12,500
Chlordiazepoxide 300 β-Phenylethylamine 75,000
Secobarbital 100,000 Desipramine 200
Clobazam 48 METHAMPHETAMINE
Norketamine 50,000 Doxepin 2,000
Clorazepate 97 d-Methamphetamine 1,000
METHADONE Imipramine 400
English 2
 of alcohol, the reaction pad will rapidly change colors beginning the test. This includes non‐alcoholic drinks,
depending on the concentration of alcohol present. This tobacco products, coffee, breath mints, food, etc.
color change is proportional to the concentration of alcohol 1. Bring the pouch to room temperature before
in the saliva specimen. By comparing with the color blocks opening it. Remove the test strip from the sealed
on the color chart printed on the pouch, an approximate pouch and use it as soon as possible after observing
blood alcohol concentration (BAC) can be determined. the reaction pad on the end of the test strip. The
Saliva Alcohol Test Strip (Saliva)
reaction pad should have a light cream color. Do
MATERIALS not use the test strip if the reaction pad has a blue
A rapid saliva test strip for the determination of MATERIALS PROVIDED color before the saliva specimen is applied or is
relative blood alcohol concentration. • Test Strips (contains Tetramethylbenzidine, otherwise discolored.
For in vitro diagnostic use only. Alcohol Oxidase (EC 1.1.3.13), Peroxidase (EC 2. Saturate the reaction pad with saliva from the
1.11.1.7) and other additives). collection cup or by applying saliva directly to the
• Collection cups reaction pad. Saturating the reaction pad usually
• Package Insert takes 6‐8 seconds. Start the timer immediately
INTENDED USE after saturating the reaction pad with saliva.
Atlas Saliva Alcohol Test Strip (Saliva) is a rapid, highly 3. Note: For hygienic and sanitary reasons, placing
MATERIALS NEEDED BUT NOT PROVIDED
sensitive method to detect the presence of alcohol in saliva
• Timer the test strip in the mouth is not recommended.
and provide an approximation of relative blood alcohol 4. Read results at 2 minutes by visually comparing
concentration (BAC) at 0.02% or greater. the color of the reaction pad to the corresponding
PRECAUTIONS
This test provides a preliminary result only. A more specific color blocks printed on the pouch to determine
• For medical or other professional in vitro
alternate chemical method must be used in order to obtain the relative blood alcohol concentration. Do not
diagnostic use only. Do not use after the
a confirmed analytical result. Gas chromatography (GC) is interpret the result after 3 minutes.
expiration date.
the preferred confirmatory method. Clinical consideration
and professional judgment should be applied to any result, • All specimens and test materials that have been
particularly when preliminary positive results are indicated. exposed to saliva should be treated as potentially
infectious.
INTRODUCTION • The used test strip and test materials should be
Two‐thirds of all adults drink alcohol.¹ It has been well discarded according to local regulations.
established that the concentration of alcohol in saliva is
comparable to that in blood.2,3 The blood alcohol STORAGE AND STABILITY
concentration at which a person becomes impaired varies • Store as packaged in the sealed pouch either at INTERPRETATION OF RESULTS
depending upon the individual. Each individual has specific room temperature or refrigerated (2‐27°C). NEGATIVE:
parameters such as size, weight, eating habits and alcohol • The test strip is stable through the expiration date No color change appears on the reaction pad. The color
tolerance that affect the level of impairment. Inappropriate printed on the sealed pouch. should match the color block on the pouch corresponding
consumption of alcohol can be a contributing factor to • The test strip must remain in the sealed pouch to a negative (‐) result. This indicates that alcohol has not
many accidents, injuries, and medical conditions. until use. been detected.
Atlas Saliva Alcohol Test Strip (Saliva) is a rapid saliva test • Do not freeze. POSITIVE:
that can be performed without the use of an instrument. • Do not use beyond the expiration date. A color change appears on the reaction pad. The BAC will
The test utilizes an enzyme system to detect alcohol in range from 0.02% to 0.30%, with the color on the reaction
saliva at blood alcohol concentrations of 0.02% or greater. SPECIMEN COLLECTION AND PREPARATION pad varying from a light blue to a dark blue, falling on or
Saliva specimens may be stored in a sealed container at between the corresponding color blocks on the pouch.
PRINCIPLE 15‐27°C for up to 4 hours prior to testing. Specimens NOTE:
Atlas Saliva Alcohol Test Strip (Saliva) is a chemical assay may be refrigerated and stored at 2‐8°C. Do not freeze The test strip is very sensitive to the presence of alcohol. A
based on an alcohol‐sensitive enzymatic reaction. Alcohol, if saliva specimens. Refrigerated specimens should be blue color lighter than the 0.02% color pad should be
present in the saliva specimen, reacts with chemicals on the brought to room temperature before testing. interpreted as alcohol positive in saliva but less than 0.02%
reaction pad and causes a color change. concentration of alcohol in blood.
Atlas Saliva Alcohol Test Strip (Saliva) consists of a plastic PROCEDURE INVALID:
strip with a reaction pad attached at the tip. The reaction Allow the test strip, specimens and/or controls to The outer edges of the reaction pad produce a slight color
pad employs a solid‐phase chemistry system which uses a equilibrate to room temperature (15‐27°C) prior to testing. but the majority of the reaction pad remains colorless.
highly specific enzyme reaction. On contact with solutions Do not place anything in the mouth for 15 minutes before Repeat the test with a new test strip, ensuring complete
saturation of the reaction pad with saliva. If the problem
persists, discontinue using the lot immediately and contact as cold medicines, breath sprays and
your local distributor. mouthwashes can produce positive results. Wait
at least 20 minutes after ingesting any such
QUALITY CONTROL products before using the test strip.
• Atlas Saliva Alcohol Test Strip (Saliva) may be
qualitatively verified by using a test solution PERFORMANCE CHARACTERISTICS
prepared by adding 5 drops of 80 proof distilled The detection range of Atlas Saliva Alcohol Test Strip
spirits to 30mL of water. This solution should (Saliva) is from 0.02% to 0.30% for the approximate relative
produce a color change on the reaction pad blood alcohol concentration. The appropriate limit for
corresponding to a 0.02% or greater BAC. The determining sobriety varies depending on local regulations.
color reaction with alcohol in saliva is somewhat Specificity
slower and less intense than with alcohol in an The Saliva Alcohol Test Strip (Saliva) will react with methyl,
aqueous solution. ethyl and allyl alcohols.
• Do not perform the control test with undiluted Interfering Substances
alcohol, as pure alcohol solutions will not produce The following substances may interfere with the Saliva
a positive result. Alcohol Test Strip (Saliva). These substances do not
normally appear in sufficient quantity in saliva to interfere
LIMITATIONS with the test.
• Atlas Saliva Alcohol Test Strip (Saliva) provides Peroxidases Mercaptans Bilirubin
only a preliminary result for the relative BAC. A Strong oxidizers Tosylates L‐dopa
secondary analytical method must be used to Ascorbic acid Oxalic acid L‐methyldopa
obtain a confirmed result. Gas chromatography Tannic acid Uric acid Methampyrone
(GC) is the preferred confirmatory method. Pyrogallol
• Failure to wait 15 minutes after smoking or
placing food, drink, or other non‐alcoholic REFERENCES
materials in the mouth before performing the test 1. Volpicellim, Joseph R., M.D., Ph.D.: Alcohol
can produce erroneous results due to possible Dependence: Diagnosis, Clinical Aspects and
contamination of the saliva by interfering Biopsychosocial Causes, Substance Abuse Library,
substances. University of Pennsylvania, 1997.
• Interpretation of visual results is dependent on 2. Jones, A.W.: Inter‐and intra individual variations in
several factors: the variability of color perception, the saliva/blood alcohol ratio during ethanol
the presence or absence of inhibitory factors, and metabolism in man, Clin. Chem. 25,1394‐1398,
the lighting conditions when the strip is read. 1979.
Caution should be taken when interpreting test 3. McColl K.E., Whiting, B., Moore, M.R. and
results due to the subjective nature of the test. Goldberg, A.: Correlation of ethanol
• Atlas test strip should not be used to determine concentrations in blood and saliva, Clin.Sci., 56,
the presence of alcohol in beverages, in undiluted 283‐286, 1979.
alcohol, or in other liquid solutions.
• Atlas test strip is highly sensitive to the presence Atlas Medical
of alcohol. Alcohol vapors in the air are William James House,
sometimes detected by the test strip. Alcohol Cowley Road, Cambridge, CB4 0WX, UK
vapors are present in many institutions and Tel: ++44 (0) 1223 858 910
homes. Alcohol is a component in many Fax: ++44 (0) 1223 858 524
household products such as disinfectant,
deodorizers, perfumes, and glass cleaners. If the PPI649A01
presence of alcohol vapors is suspected, the test Rev B (21.02.2010)
should be performed in an area known to be free
of vapors.
• Ingestion or general use of over‐the‐counter
medications and products containing alcohol such
CCHM2LAB W14 • Saliva: 30mL polyethylene bottle (plastic)
Blood: 10mL plain test tube for serum (non-additive: red top)
Drug Test Specimen Collection •
• Hair: self-sealed transparent plastic bag
Sweat: BFAD approved sweat patch
and Accessioning
•
• Tissues: screw capped plastic container
TYPES OF SPECIMEN COLLECTION
Drug Test Specimen Collection a) Single specimen collection – specimen is entirely placed in a
Drug Testing single 60mL bottle
❑ The best practices for drug testing in the Philippines are based on b) Split specimen collection – is collected at same time but placed
the general principles that have been established internationally. in 2 separate containers at least 30mL each
They are designed to ensure that the entire drug testing process
is conducted to give accurate and reliable information about TYPES OF SPECIMEN
client/donor/subject’s drug use, as mandated by Republic Act 1. Urine 4. Saliva
9165 otherwise known as the “Comprehensive Dangerous Act 2. Hair 5. Sweat
of 2002”. 3. Blood 6. Tissue
❑ The East Avenue Medical Center is designated as the National Urine Hair specimen
Reference Laboratory (NRL) for Environmental and Occupational  MOST COMMON  Secondary specimen of choice
Health, Toxicology and Micronutrient Assay by virtue of  Least expensive  Drugs are absorbed through hair
Department Order No. 393-E s. 2000.  Easy to do shaft
 Standardized  Expensive and tedious
Chain of Custody and Accessioning procedures  2x more sensitive than urine test
Chain of Custody  Detects use within the  Do not detect recent use
A laboratory shall use documented chain of custody week  Detects chronic substance abuse
procedures to maintain control and accountability of specimens. The  Abstaining can produce  Requires 1.5 x 1.5 cm hair clump
date and purpose shall be recorded on an appropriate Custody and negative reaction  Not affected by drug abstinence
Control Form each time a specimen is handled or transferred and  Established specimen  Can determine temporal pattern
every individual in the chain shall be identified. validity tests
Blood Saliva
Accessioning
 MOST ACCURATE  Uncommon method
1. Provide a unique accession number upon entry of the specimen to
 Least common method  Easy to administer
the laboratory;
 Short detection time  Short detection time
2. Inspect the specimen submitted and the CCF to verify the integrity
 Measures concentration  No reference standard
and identity of the specimen;
of compound directly
3. Examine the packaging for evidence of tampering in transit;
Sweat (patch) specimen
4. Compare the information on the sample bottles within the
package; and  Requires wearing of patch 1-2 weeks
5. Document all discrepancies.  Uncommon methods
 No reference standards develop
 Surface contaminations can cause FALSE POSITIVE
 Can detect use for extended period of time

TYPES OF SPECIMEN FOR COLLECTION

SPECIMEN REASON FOR MINIMUM STORAGE
COLLECTION QUANTITY
URINE Pre-employment, 60mL (single) Prolonged
random, reasonable 30mL each for storage at -20C
suspicion/cause, split sample
Collection Site Requirements mandatory
✓ A toilet/stall SALIVA Pre-employment, 2mL (single) Deep-frozen at
✓ Handwashing facility outside of toilet (ORAL random, reasonable 1.5 and 0.5 least -8 to -10C
✓ Specimen booth FLUID) suspicion/cause (split)
✓ Toilet coloring reagent

SCALP Pre-employment, 100mg or Stored at cool

HAIR random, return to equivalent and dry place
On Site Specimen Collection (beard, duty, follow up, number of
− Specimen are collected at a designated area within the drug stash) suspicion to actual stands
testing facility test
− For all mandatory tests EXCEPT for crime scene and post- Sweat DOH cleared
accident. patch worn for
Remote Collection 7-14 days
− Specimen are collected at temporary facility located at a remote Blood 5mL Separate serum
site. & immediate
Allowable Conditions: freeze @ -20C
• Workplace, school, jail, prison or rehab centers Tissue Macerated &
o Random frozen
o Follow-up
o Reasonable suspicion Reasons for Drug Testing:
o Cause » Mandatory tests – for firearm license, law enforcement field, public
• Critically ill or disabled officials, criminal case
• Secure permit from BHFS or CHD for remote collection 10 » Random tests – school, workplace
working days prior to scheduled collection » Reasonable suspicion/cause – personal issues, etc.
• No testing/examination to be done at remote collection site » Post accidents
» Follow up, return to duty – rehab, suspension of work due to
Collection Supplies: (Specimen containers) usage
• Urine: 30 or 60mL, polyethylene bottle wide mouth with screw » Pre-employment
cap (plastic)
Methods of Specimen Collection: Judicial proceedings or upon Up to a year
1) Observed Collection (in the presence of ASC) request
2) Unobserved collection (in the absence of ASC) Specimen Rejection at Collection Site:
3) Submitted samples (subjected to visual observation) All rejected specimen shall be documented and reported to the Head
4) Subject to validity tests of the Laboratory (HOL)
*ASC – Authorized Specimen Collector Memorandum for Record (MFR)
Preliminary Procedures Prior to Collection • A record to document that recovery, corrective and remedial
1. Verify identity of donor measures to administrative errors
2. Explain collection procedure • Accomplished by ASC and other lab personnel
3. Answer questions regarding the procedure • Errors are not corrected by an MFR shall be rejected or
cancelled
Donor identification:
▪ Photo ID (driver’s license, employee ID, passport) The SPECIMEN COLLECTION is considered the “WEAKEST
▪ Identification by authorized agency representative LINK” of the drug testing program. All efforts must be done to make the
▪ Any other ID allowed by agency’s workplace drug testing plan collection legally, forensically, and technically indefensible.

STEPS IN COLLECTION: Information that should be present in the container:

1. Check on supplies, security of collection area 1. Urine: name, age, gender, sex, sample volume, date & time of
2. Check of ID of donor collection, signature of ASC and client, storage of sample
3. Explains/answers about collection procedure 2. Saliva: same w/ urine
4. Fills up step 1 of CCF 3. Hair: name, age, sex + weight of sample, date & time collected,
5. Gives specimen bottle or ask donor to select specimen bottle from signature of ASC and client
available supplies. Labels legibly.

ASC – observes collection one at a time

▪ Close attention to collection
▪ Observe unusual behavior, if present, repeat under DIRECT
OBSERVED COLLECTION
▪ Collector performs the ff:
o Asks donor to remove outer garments (coat)
o Examinees pockets etc. for presence of adulterants
o Asks donor to wash and dry hands
▪ Measures temperature, volume, inspect for adulteration and
substitution. Fills up Step 2 and initiates Step 3 of CCF
▪ Closes, places and initials seals over the lid bottle in front of donor
▪ Asks donor to fill up and sign Step 5

FORMS OF TAMPERING SPECIMEN:

A. Dilution:
• Internal dilution:
= Drinking plenty of water
= Diuretic like Lasix, tea, coffee, beer
• External dilution:
= Addition of water to specimen
B. Substitution
C. Adulteration
Additives/Adulterants:
Bleach Table salt
Ammonia Vinegar
Liquid soap Visine eye drops

Custody and Control Forms (CCF)

→ A form used to document the security of specimen
o All steps of collection
o Persons who handles the specimen
o Status and integrity of specimen
o Pertinent info
→ A form document chain of custody from collection to transport,
analysis and releasing of reports
→ Improper entries will invalidate testing procedure
→ With legal and forensic implications
→ 3 copies (patient/employer, ASC, lab)

DATA ENTRIES IN CCF

 Patient info: name, address, sex, etc.
 Names, signature of handling specimen
 Status of specimen: temp, physical appearance, other remarks
 Drug tests requested (meth, THC are common)
 Result of test (screening/confirmatory/NRL level)

SPECIMEN RETENTION:
Negative result 5 days
Positive result Min. of 5 days
Adulterated, substituted, invalid 15 days
result
CCHM2LAB W15 − Less than 1% is excreted unaltered in

Drug Testing Cocaine

the urine for 3-7 days.
− Self-administered through nasal
inhalation, intravenous injection and
Methods for Detection of Substances of Abuse free base smoking.
 Enzymatic – solely dependent on presence of particular compounds, − It has a biologic half-life of 0.5-1.5
(detecting several compounds w/ same characteristics) hours.
Ex. Saliva Alcohol Breath Test → alcohol is measured by − It is a potent central nervous system
assuming the possible BAC based on the saliva spx involving stimulant and a local anesthetic.
enzymes, w/c deoxidation of products to induce change in color − Brings extreme energy, restless,
involving tetramethylbenzidine. tremors, oversensitivity and spasms.
 Capillary Electrophoresis – differentiate selectivity of analytes based − Large amounts: fever,
on different physicochemical principles of separation. unresponsiveness, difficulty in
• Variation of the TLC, where it has the advantages same as the breathing, unconsciousness.
High-performance liquid chromatography. − Benzoylecgonine → detected for 24-
 Homogenous Immunoassay – solution does not need to undergo 48 hours after exposure, with a
separation biologic half-life of 5-8 hours exceed
a) EMIT (Enzyme Multiplied Immunologic Technique) – uses 300 ng/mL.
enzyme labeled drug that competes with the drug in the sample. Methylenedioxy- − Designer drug first synthesized in
→ Involves immunologic type of rxn in w/c the active site methamphetamine 1914 by a German drug company for
of the enzyme will be blocked w/ the anti-drug antibody (Ecstasy) the treatment of obesity.
w/c will result the decrease of enzymatic activity. The − Produces perceptual changes such as
free drug present in the analyte of the serum/urine increased sensitivity to light, difficulty
sample will compete w/ the antibody-drug enzyme in focusing, clenching of jaws and
complex separating it further. blurred vision in some users.
 Chromatographic Methods − Not stimulant but effect is same as
a) Thin-Layer Chromatography (TLC) – used for any type of amphetamine drug.
sample (serum/urine/gastric fluid) − Increases blood rate and pressure and
→ The extraction of the specific drug will be separated heart rate of an individual
would be pH-dependent, that you could determine a − Mechanism of action is due to release
particular group acc. to w/c pH it will be extracted from of the neurotransmitter, serotonin and
such as acidic drugs (pH: 4.5) e.g barbiturates; dopamine
alkaline drug (pH: 9.0) e.g. opiates Methadone − Narcotic pain reliever for medium to
b) Liquid Chromatography - Mass Spectroscopy (LC-MS) – severe pain - last between 12 to 48
for detection of poisons in acute or chronic intoxication, hours
therapeutic drug identification and quantitation, − Used in the treatment of Heroin /
pharmacokinetics and drug metabolism studies Opiate dependence: Vicodin,
→ For confirming (+) test results from a screening assay Percocet, Morphine, etc.
for non-volatile compounds but is NOT the gold − Currently an ACCEPTABLE method for
standard for drug abuse. detoxification of px & undergoing
c) High Performance Liquid Chromatography (HPLC) therapy (rehab)
→ Alternative to GC-MS, but GC-MS is preferred for Morphine − Opioid analgesics which control pain
confirmatory testing. MAJOR metabolic by depressing the central nervous
product of codeine system
→ Allows quantitative measurement of drugs as well as
and heroin − Produces higher tolerance levels and
separation of same drug.
→ For tricyclic antidepressants and its metabolites. physiological dependency in users and
d) Gas Chromatography may lead to substance abuse.
i. Gas-Liquid Chromatography (GLC) − Excreted unmetabolized in urine
 Legally accepted method for ethanol testing. − Detectable in the urine for several
ii. Gas Chromatography with Infrared Spectroscopy days after an opiate dose
 Used for detection of amphetamines. Methamphetamine − Increases mental alertness and
iii. Gas Chromatography-Mass Spectroscopy (GC-MS) Active reagent of physical capacity and has anorectic
 GOLD STANDARD for confirmation of screening shabu (most common) property.
methods such as TLC and EMIT − Acute psychotic syndromes: auditory
 Allows for detection of low levels of drugs, very and visual hallucinations, paranoia
sensitive and specific for different drugs of abuse − Toxic Effects: Palpitations,
as well as its metabolites. hypertension, cardiac arrhythmias,
convulsions, pancytopenia, mental
Barbiturates − Barbiturates are sedatives, hypnotics, impairment, teeth grinding
anti-convulsants, central nervous Tetrahydrocannabinol − MOST POTENT component or the
system depressants. (most common) psychoactive substance of marijuana.
− Barbiturates taken at 400 mg/day for − Induces a sense of well-being and
2-3 months produce significant euphoria
dependence. − detected in urine for 3-5 days up to 4
− Abstinence/withdrawal → death weeks for chronic users.
− Barbiturates are unaltered in the urine − Principal psychoactive agent: delta-9-
and could be detected for 4-7 days. tetrahydrocannabinol
Benzodiazepines − Clinically used as sedative for surgical − Urinary metabolite: 11-nor-
and medical procedures, treatment for deltatetrahydrocannabinol (THC-
anxiety and sleep disorders. COOH)
− Withdrawal Symptoms → trouble in → detected in drug testing kit
sleeping, gastrointestinal upset, feeling
unwell, loss of appetite, sweating,
trembling, weakness, anxiety, and
changes in perception.
Test Kits: INTERPRETATION OF RESULTS
» May detect same metabolites px has been getting from medications. NEGATIVE: Two lines appear. One colored line should be in the control
» Based on the principle of immunoassay specifically Competitive region (C), and another apparent colored or faded color line adjacent
binding (Ag-Ab reaction) present in the gold colloid reaction pad should be in the test region (T). This negative result indicates that the drug
» POSITIVE: absence of the changes in color for the test area. concentration is below the detectable level.
» TESTING:
• Urine specimen will migrate upwards through capillary action (in POSITIVE: One colored line appears in the control region (C) (bcos the
the calibrator) drugs will saturate Ab). No line appears in the test region (T). This positive
• If drug is present w/c is beyond the “cut-off”, it will then produce result indicates that the drug concentration is above the detectable level.
reaction color. But if below, it will not be able to saturate the
binding site. INVALID: Control line FAILS to appear. Insufficient specimen volume or
• Calibrator: specific analyte being tested. incorrect procedural techniques are the most likely reasons for control
• Cut-off: expressed in nanogram (ng)/mL; the actual amount that line failure. Review the procedure and repeat the test using a new test
must be reached before it can give significant effect to the panel. If the problem persists, discontinue using the lot immediately and
competitive binding. contact your local distributor.
» REACTION:
• If drug is not present, or calibrator is present but below the cut-off INDEX OF SYMBOLS
value → no saturation on the binding site → no antigen → antibody
will react to the drug protein conjugate and visible will line up.
• If present, the drugs beyond the cut-off value will saturate the
binding sites of the Ab → the metabolite will bind to the Ab → no
available Ab w/c will react to the test line region → no colored
line.
• Test line: contains the drug Ab-coupled particles w/
corresponding drug conjugate Storage: Can be refrigerated or at room temp. DO NOT FREEZE.
• Control line: contains goat Ab, ensuring acceptability and If refrigerated, always put in room temp first.
sufficiency. Produces color whether its negative/positive if sample
is adequate in amount.

Single test kit

Multi test kit

Deep stick

TEST PROCEDURE:
1. Allow the test panel and urine specimen to equilibrate to room
temperature (15-30˚C) prior to testing.
2. Bring the pouch to room temperature before opening it. Remove the
test panel from the sealed pouch and use it as soon as possible.
(check for expiration date, sealed, untampered)
3. Take off the cap outside of the test end.
4. With arrow (s) pointing toward the urine specimen, immerse the test
panel vertically in the urine specimen for at least 10-15 seconds (Dip
stick) 3 drops/3 gtts (well) approx. 100 uL
5. Do not pass the arrows on the test panel when immersing the panel.
The same procedure is followed for a multi-panel test card.
6. Place the test panel on a non-absorbent flat surface, start the timer
and wait for the colored line (s) to appear. The results should be read
at 5 minutes. DO NOT interpret results after 10 minutes.