CupScan Test Procedure
- Remove the CupScan from the sealed foil pouch. If collected urine sample has been refrigerated, bring to room temperature (15°C-28°C)
- Remove the lid and collect the sample, ensuring that the sample is above the minimum fill line.
- Secure the lid tightly and place the CupScan on a flat surface.
- Remove the key from the lid and insert it in the side chamber, push to release the sample into the test zone.
- Place the CupScan on a flat surface
- Remove the result window cover to view the results. Results should be viewed at 5-8 minutes.
Do not interpret the results after 10 minutes.
Interpretation of Results
Negative: The appearance of a Control line (C) in all six of the test windows and a Test line (T) for
a specific drug indicates a negative result. The colour intensities of the Control line and specific drug
line may not be the same. The negative result does not indicate the absence of drug in the specimen;
it only indicates that the concentration of the specific drug in the specimen is less than its cut-off
level.
Positive: The appearance of only a Control line (C) in any of the six test windows and no Test line
(T) next to a specific drug indicates that the test result is positive for that drug. This is an indication
that the specific drug in the specimen is above its cut-off level.
Invalid: A distinct Control line (C) in all six of the test windows should always appear. If there is
no line formation in the C position, the test result is invalid. Retest the sample with a new device.
Note: A very faint line for a specific drug in the result window does not indicate that the amount
of drugs in the sample is near or above the cut off level. A faint line is still indicative of a negative
result.
When reading the results on CupScan, a line next to the numbers 1 or 2 indicates a negative result for
the corresponding drug. The absence of a line next to the numbers 1 or 2 indicates
a positive result for the corresponding drug.
Alcohol
Alcohol intoxication can lead to
loss of alertness, coma and death as well as
birth defects. The BAC at which a person
becomes impaired is variable. The United
States Department of Transportation (DOT)
has established a BAC of 0.02% (0.02g/dL)
as the cut-off level at which an individual
is considered positive for the presence
of alcohol.
Determination of ethyl alcohol in blood,
saliva and urine is commonly used for
measuring legal impairment, alcohol
poisoning, etc. Gas chromatography
techniques and enzymatic methods
are commercially available for the
determination of ethyl alcohol in human
fluids. The test is designed as a screening
tool to rapidly determine if the alcohol level
is higher than 0.04%.
Principles of the Tests
Urine Alcohol
The Alcohol Test is based on the high specify
of alcohol oxidase (ALOx) for ethyl alcohol
in the presence of peroxides and enzyme
substrate such as tetramethylbenzidine
(TMB) as shown in the following:
ALOxPeroxidase
EtOH+TMB"CH3CHO+Coloured TMB
The distinct colour on reactive pad could
be observed in less than 1 minute after the
tip was contacted with urine samples with
the ethyl alcohol concentration greater than
0.04%. It should be pointed out that other
alcohols such as methyl, propanyl and allyl
alcohol would develop the similar colour on
the reactive pad. However, these alcohols
are not normally present in urine.
Adulteration Test
The adulteration tests are built-in strips
to help to verify the integrity of the urine
specimen to be tested. Only fresh and
uncentrifuged urine samples without
preservatives should be used.
In general, all adulteration tests are based
on the chemical reactions of the indicator
reagents on the pads with components in
the urine sample effecting colour changes.
Results are obtained by comparing the
colour on each of the test pads with the
corresponding pad on the colour chart
provided.
Creatinine: Testing for sample dilution
In this assay, creatinine reacts with a
creatinine indicator in an alkaline condition
to form a purplish- brown colour complex.
The concentration of creatinine is directly
proportional to the colour intensity of the
test pad.
pH: Testing for the presence of acidic
or alkaline adulterants. This test is based
on the well-known double pH indicator
method that gives distinguishable colours
over a wide pH range. The colours range
from orange (low pH) to yellow and green
to blue (high pH).
Adulteration Test &
Urine Alcohol Test
Compare the reactive pad with this coloured chart:
Note: Urine Alcohol is only available on Cupscan II
Semi quantitative results are obtained by visually comparing the colour of each pad with
the corresponding test colour chart provided below. A borderline (+/-) result in the test
zone should be considered a negative result.
Adulteration Test
For best results, performance of the
adulteration test should be confirmed
by testing known negative and positive
specimens or controls whenever a new test
is performed.
Creatinine: Daily creatinine excretion,
related to muscle mass of the human body,
is usually constant. The DOT guideline
states that urine specimens with creatinine
levels of less than 20mg/dl are indications
of adulteration. Although these ranges are
affected by age, sex, diet, muscle mass and
local population distribution, samples with
creatinine levels lower than 20mg/dl should
be considered adulterated.
pH: Normal urine pH ranges from 4.5 to
8.0. Values below pH 4.0 or above pH 9.0
are indicative of adulteration.
Drug Groups Tested For:
Amphetamine (AMP)
Amphetamines are a class of potent sympathomimetic agents with therapeutic applications. The most
common Amphetamines are d-Amphetamine and d,l-Amphetamine. Amphetamines are central nervous
system stimulants that cause the neurotransmitters epinephrine, norepinephrine and dopamine to be released
into the brain and body giving users a feeling of euphoria, alertness, and increased energy. Chronic abuse of
Amphetamine leads to tolerance and a drug reinforcement effect. Cardiovascular responses to Amphetamine
include increased blood pressure and cardiac arrhythmia. More acute responses produce anxiety, paranoia,
hallucinations and psychotic behavior. Amphetamine is metabolized by a number of pathways. In general,
acid urine promotes excretion whereas alkaline urine retards it. In 24 hours, approximately 79% of the
Amphetamine dose is excreted in acid urine and about 45% in alkaline urine. Typically, about 20% is
excreted as unchanged Amphetamine. Unchanged Amphetamine can be detected up to 10 days after use.
Benzodiazepines (BZO)
Benzodiazepines are a class of widely prescribed central nervous system depressants, which have anxiolytic,
hypnotic, anticonvulsant and muscle relaxant effects. Chronic abuse can result in addiction and tardive
dyskinesia (involuntary twitching). Acute higher doses lead to drowsiness, dizziness, muscle relaxation,
lethargy, coma and possible death. The effects of Benzodiazepine use lasts between 4 – 8 hours. Many of
the Benzodiazepines share a common metabolic route and are excreted as Oxazepam in urine. Oxazepam is
detectable in the urine for up to 7 days after drug use.
Cocaine (COC)
Derived from the leaves of the cocoa plant, cocaine is a potent central nervous system stimulant as well as
a local anesthetic. Some of the psychological effects induced by cocaine are: euphoria, confidence and a
sense of increased energy, accompanied by increased heart rate, dilation of the pupils, fever, tremors and
sweating. Continued ingestion of cocaine could induce tolerances and physiological dependency, which
leads to its abuse. Cocaine is consumed by smoking, intravenous, intranasal or oral administration and is
excreted in the urine primarily as benzoylecgonine in a short period. Benzoylecgonine has a biological halflife
of between 5 – 8 hours, which is much longer than that of cocaine (0.5 – 1.5 hours), and can be generally
detected after cocaine use or exposure for up to 3 days.
Methamphetamine (MET)
Methamphetamine is the most popular synthetic derivative of the Amphetamines. It is a potent
sympathomimetic agent with therapeutic applications. Large doses will lead to enhanced stimulation of the
central nervous system and can induce euphoria, alertness, reduced appetite, and a sense of increased energy
and power. More acute responses produce anxiety, paranoia, psychotic behavior and cardiac dysrhythmias.
Methamphetamine is excreted in the urine as Amphetamine and oxidised and deaminated derivatives.
However, 10 - 40% of Methamphetamine is excreted unchanged. Methamphetamine is generally detectable
in the urine for 3 to 5 days after use.
Opiates (OPI)
Opioid analgesics are comprised of a large group of substances that control pain by depressing the central
nervous system. Acute, high doses used by abusers or addicts can cause depressed coordination, disrupted
decision making, decreased respiration, hypothermia and coma. Morphine is excreted unmetabolized and is
the marker metabolic product of Opiates. Morphine and morphine glucuronide is detectable in urine several
days after an Opiates dose.
Marijuana (THC)
The agents of Marijuana that cause various biological effects in humans are called cannabinoids.
Cannaboids are a central nervous system stimulant that alters mood and sensory perceptions, produces
loss of coordination, impairs short-term memory and produces symptoms of anxiety, paranoia, depression,
confusion, hallucination and increased heart rate. Large doses of cannabinoids could cause the development
of tolerances and physiological dependency and lead to abuse. A tolerance to the cardiac and psychotropic
effects can occur and the withdrawal syndrome produces restlessness, insomnia, anorexia and nausea. THC
is the primary active ingredient in cannabinoids. The main metabolite excreted in the urine is 11-nor-r9-
THC-9-COOH, which is found within hours of exposure and remains detectable in the urine between 10-30
days after smoking and even longer for chronic smokers.
Principle of Test
The CupScan test is based on the principle of solid-phase immunoassay technology to detect the
presence of any of the six drugs (AMP, BZO, COC, MET, OPI and THC), and/or their immunoreactive
metabolites in urine. The assay relies on the competition for binding antibodies between drug
conjugate and free drug, which may be present in the urine specimen being tested. When a drug is
present in the urine specimen, it competes with the drug conjugate for the limited amount of antibodydye
conjugate. When the amount of drug is equal or more than the cut-off levels of any of the drugs,
it will prevent the binding of the drug conjugate to the antibody. Therefore, a positive urine specimen
will not show a coloured band on the test line zone, indicating a positive result, while the presence
of a coloured band indicates a negative result. A control line is present in the test window to work
as a procedural control. This coloured band should always appear on the control line zone if the test
device is stored in good condition and the test is performed appropriately. The control line serves to
validate the test results.
Storage and Stability
The CupScan test device should be stored at 2° to 30° C and will be effective until the expiration date.
Specimen Collection and Preparation
A minimum volume of sample urine is required for each test which is indicated by the minimum
fill volume line indicated on the cup. Fresh urine specimens do not need any special handling or
treatment. If the assay is not performed immediately, the urine specimen may be refrigerated at 2 - 8’C
for up to 7 days. Refrigerated specimens should be brought to room temperature before testing. Urine
specimens exhibiting a large amount of precipitate or turbidity should be centrifuged or allowed to
settle before testing.
Precautions
- For in vitro diagnostic and forensic use only.
- Do not use the product beyond the expiration date.
- Handle all specimens as potentially infectious.
- Do not open foil pouch until it is ready to be tested.
Quality Control
- The control band is an internal reagent and procedural control. It will appear in each of the six
test windows if the test has been performed correctly and the reagents are reactive.
- Control standards can be used to validate reagent performance and establish test reliability.
Controls, which are not provided with this test, are commercially available.
Expected Results
The CupScan Test is a qualitative assay. The amount of drug and metabolites present in the urine
cannot be estimated by the assay. The test is intended to distinguish negative results from presumptive
positive results above the cutoff concentrations. All positive results must be confirmed using an
alternate method, preferably GC/MS (as outlined in the AS/NZS 4308 standards).
Intended Use
CupScan is a rapid, one step immunochromatographic assay for the qualitative determination of
Amphetamine (AMP), Benzodiazepines (BZO), Cocaine (COC), Methamphetamine (MET), Opiates
(OPI) and Marijuana (THC), and/or their metabolites in human urine.
CupScan provides only presumptive results and as per the Australian/New Zealand standards
should be confirmed by more specific alternate chemical methods such as gas chromatography/mass
spectrophotometry (GC/MS). Positive results should be justified with compelling clinical evidence
and professional judgement. CupScan is not designed to monitor drug levels, but only to screen urine
for the presence of the six Drugs of Abuse (DOA).
The CupScan Test detects the major metabolites of the six drugs at the following cutoff levels based
on the recommendations in the Australian/New Zealand Standards AS/NZS 4308 for screening of the
following drugs in urine:-
| Amphetamine |
300 µg/L |
| Benzodiazepines |
200 µg/L |
| Cocaine Metabolites |
300 µg/L |
| Methamphetamine |
300 µg/L |
| Opiates |
300 µg/L |
| Cannabis Metabolites |
50 µg/L |
Performance Characteristics
Sensitivity
The following table lists compounds
that are detected by CupScan tests.
Compounds
AMP
- d-Amphetamine
- d,l-Amphetamine
- l-Amphetamine
- (±)3,4-Methylenedioxyamphetamine
BZO
- Nitrazepam
- Chloradiazepoxide HCl
- Clobazam
- Desmethyldiazepam
- Oxazepam
- Temazepam
- Alprazolam
- Bromazepam
- Diazepam
- Flunitrazepam
- Lorazepam
- Clonazepam
- Flurazepam
|
COC
MET
- (+)Methamphetamine
- (±)3,4-Methylenedioxymethamphetamine
OPI
- Morphine
- Morphine-3-ß-glucuronide
- Codeine
- Hydromorphone
- Nalorphine
- Heroin
- Hydrocodone
- Normorphine
- Naloxone
- Natrexone
|
THC
- 11-nor-r9-THC-9-COOH
- 11-nor-r8-THC-9-COOH
- 11-hydroxy-r9-THC
- r8-Tetrahydrocannabinol
- r9-Tetrahydrocannabinol
|
Interference testing
The following substances did not
interfere with the CupScan test.
- Glucose
- Human albumin
- Human haemaglobin
- Urea
- Uric acid
References
- Urine Testing for Drugs of Abuse, NIDA Research Monograph 73, (1986).
- Critical Issues in Urinalysis of Abused Substances: Report of the Substance Abuse Testing committee, clinical Chemistry, 34(3), 617 (1988)
- Greenblatt DJ, shader RI: Benzodiazepines in Clinical Practice. New York: Raven Press, 1974
- Stewart DJ et al: Cocaine and norcocaine hydrolysis by liver and serum esterase. Clin. Pharmacol ther 1978 25:464-468
- Blum, K., Handbook of Abusable drugs, Gardener Press, Inc., New York, NY, 1st Ed., (1984).
- Baselt RC., Disposition of Toxic Drugs and Chemicals in Man, 3rd Ed.,Chicago, IL. Year Book Medical Publishers Inc., 780-783, (1990).
- Mandatory Guidelines for Federal Workplace drug Testing Programs, Fed. Reg. 53(69):11970-89 (1988).
- Procedures for the collection, detection and quantitation of drugs of abuse in urine, Australian/New Zealand Standards AS/NZS 4308.
