One Step Cotinine Test Device
A rapid, one step test for the qualitative detection of Cotinine
(nicotine metabolite) in human urine.
For Determination of Smoking Status Only.
The COT One Step Cotinine Test Device (Urine) is a lateral flow
chromatographic immunoassay for the detection of Cotinine in human urine at a
cut-off concentration of 200 ng/mL. This test will detect other related
compounds, please refer to the Analytical Specificity table in this package
This assay provides only a preliminary
analytical test result. A more specific alternate chemical method must be used
in order to obtain a confirmed analytical result. Gas chromatography and mass
spectrometry (GC/MS) is the preferred confirmatory method. Clinical
consideration and professional judgment should be applied to any drug of abuse
test result, particularly when preliminary positive results are used.
Cotinine is the first-stage metabolite of nicotine, a toxic
alkaloid that produces stimulation of the autonomic ganglia and central nervous
system when in humans. Nicotine is a drug to which virtually every member of a
tobacco-smoking society is exposed whether through direct contact or second-hand
inhalation. In addition to tobacco, nicotine is also commercially
available as the active ingredient in smoking replacement therapies such as
nicotine gum, transdermal patches and nasal sprays. In a 24-hour urine,
approximately 5% of a nicotine dose is excreted as unchanged drug with 10% as
cotinine and 35% as hydroxycotinine; the concentrations of other metabolites are
believed to account for less than 5%.1 While
cotinine is thought to be an inactive metabolite, it’s elimination profile is
more stable than that of nicotine which is largely urine pH dependent. As a
result, cotinine is considered a good biological marker for determining nicotine
use. The plasma half-life of nicotine is approximately 60 minutes
following inhalation or parenteral administration.2
Nicotine and cotinine are rapidly eliminated by the kidney; the
window of detection for cotinine in urine at a cutoff level of 200 ng/mL is
expected to be up to 2-3 days after nicotine use. The COT One Step Cotinine Test
Device (Urine) is a rapid urine screening test that can be performed without the
use of an instrument. The test utilizes a monoclonal antibody to selectively
detect elevated levels of Cotinine in urine. The COT One Step Cotinine Test
Device (Urine) yields a positive result when the Cotinine in urine exceeds 200
The COT One Step
Cotinine Test Device (Urine) is an immunoassay based on the principle of
competitive binding. Drugs which may be present in the urine specimen compete
against the drug conjugate for binding sites on the antibody. During
testing, a urine specimen migrates upward by capillary action. Cotinine, if
present in the urine specimen below 200 ng/mL, will not saturate the binding
sites of antibody coated particles in the test device. The antibody coated
particles will then be captured by immobilized Cotinine conjugate and a visible
colored line will show up in the test line region. The colored line will not
form in the test line region if the
Cotinine level exceeds 200 ng/mL
because it will saturate all the binding sites of anti-Cotinine antibodies. A
drug-positive urine specimen will not generate a colored line in the test line
region because of drug competition, while a drug-negative urine specimen or a
specimen containing a drug concentration less than the cut-off will generate a
line in the test line region. To serve as a procedural control, a colored line
will always appear at the control line region indicating that that proper volume
of specimen has been added and membrane wicking has occurred.
The test device contains mouse
monoclonal anti-Cotinine antibody-coupled particles and Cotinine-protein
conjugate. A goat antibody is employed in the control line system.
• Do not use after the expiration date.
• The test device should remain in the sealed
pouch until use.
• All specimens should be considered potentially
hazardous and handled in the same manner as an infectious agent.
• The used test device should be discarded
according to local regulations.
Storage and Stabliity
Store as packaged in the sealed pouch either at room temperature or
refrigerated (2-30°C). The test device is stable through the expiration date
printed on the sealed pouch. The test device must remain in the sealed pouch
until use. DO NOT FREEZE. Do not use beyond the expiration date.
The urine specimen must be
collected in a clean and dry container. Urine collected at any time of the day
may be used. Urine specimens exhibiting visible precipitates should be
centrifuged, filtered, or allowed settle to obtain a clear supernatant for
Urine specimens may be stored at 2-8°C for up to 48 hours prior to
assay. For prolonged storage, specimens may be frozen and stored below -20°C.
Frozen specimens should be thawed and mixed before testing.
Materials Required Buy Not Provided
Specimen collection container
Directions For Use
How To Use
Cotinine Test Devices
Allow test device, urine specimen to reach room temperature
(15-30°C) prior to testing.
1.Bring the pouch to room temperature
before opening it. Remove the test device from the sealed pouch and use it as
soon as possible.
2.Place the test device on a clean and
level surface. Hold the dropper vertically and transfer 3 full drops of urine
(approx. 100 µL) to the specimen well (S) of the test device, and then start the
timer. Avoid trapping air bubbles in the specimen well (S). See the illustration
3.Wait for the colored line(s) to
appear. The result should be read at 5 minutes. It is important that the
background is clear before the result is read. Do not interpret the result after
Interpretation of Results
(Please refer to the illustration above)
NEGATIVE:* Two lines appear. One
colored line should be in the control line region (C), and another apparent
colored line should be in the test line region (T). This negative result
indicates that the Cotinine concentration is below the detectable level (200
*NOTE: The shade of color in the test line region (T) may vary, but it
should be considered negative whenever there is even a faint line.
POSITIVE: One colored line appears in the control line region (C). No
line appears in the test line region (T). This positive result indicates that
the Cotinine concentration exceeds the detectable level (200 ng/mL).
INVALID: Control line fails to appear. Insufficient specimen volume or
incorrect procedural techniques are the most likely reasons for control line
failure. Review the procedure and repeat the test using a new test device. If
the problem persists, discontinue using the lot immediately and contact your
A procedural control is included in
the test. A colored line appearing in the control line region (C) is considered
an internal procedural control. It confirms sufficient specimen volume, adequate
membrane wicking and correct procedural technique.
1.The COT One Step Cotinine Test
Device (Urine) provides only a qualitative, preliminary 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.1,2
2.It is possible that technical or
procedural errors, as well as other interfering substances in the urine specimen may cause
3.Adulterants, such as bleach and/or
alum, in urine specimens may produce erroneous results regardless of the
analytical method used. If adulteration is suspected, the test should be
repeated with another urine specimen.
4.A positive result indicates only
that the presence of Cotinine is above the cut-off concentration. It does
not indicate or measure level of consumption.
What is cotinine?
[COAT-e-neen] is a chemical that is made by the body from nicotine,
which is found in cigarette smoke. Since cotinine can be made only from
nicotine, and since nicotine enters the body with cigarette smoke,
cotinine measurements can show how much cigarette smoke enters your
As far as we
know, cotinine itself is not harmful. Cotinine is used simply to measure
how much tobacco smoke has entered your body. However, many studies show
that some of the 4,000 other chemicals found in tobacco smoke are
Why should I
have a cotinine test?
If you are
serious about stopping or reducing your smoking, or if you are
interested in the amount of smoke that has entered your body, this test
can be very useful. By knowing what your starting level of cotinine is,
you can see how successful your efforts to stop smoking are.
test is a simple
at home test that can measure cotinine in human urine.
you just ask how much I smoke? Or just keep track?
behavior varies. For example, two people could each smoke a pack of
cigarettes a day. One may smoke unfiltered cigarettes, inhaling deeply
with each puff, while the other may smoke a low tar, filtered cigarette,
puffing lightly and smoking only half of each cigarette. The cotinine
test would be able to show a difference in the amount of cigarette smoke
entering the bodies of these two smokers.
cotinine is normal?
People who do
not smoke or who are not exposed to other peoples' smoke should not have
measurable cotinine. People who do smoke will have a cotinine level of
10 or higher in their blood, and a typical smoker has levels of 150 to
450 units. Levels in urine are ten times higher.
How can I
reduce my cotinine?
The only way
to reduce your cotinine level is to stop or reduce your exposure to
should it take for me to see a drop in my cotinine level if I stop
how high your level is to begin with, your level could drop to that of a
nonsmoker in 7 to 10 days.
If I stop
smoking, then start again, how soon will cotinine show up in my body?
test can detect cotinine as soon as one day after you've had a cigarette.
If I switch
to a low nicotine cigarette, will my cotinine level drop?
It might, but
it depends on how you smoke low nicotine cigarettes. To satisfy a
craving for nicotine, some people smoke more low nicotine cigarettes
than they would regular cigarettes, and their cotinine level may
patches, gum, or aerosols have an effect on cotinine levels?
all use nicotine, these devices can increase cotinine levels. If you are
having a cotinine test, make sure that you mention on the lab slip that
you are using nicotine replacement products.
other people's smoke? Won't my cotinine level increase if I breathe
other people's smoke?
If you breathe
a lot of cigarette smoke even though you yourself don't smoke, your
cotinine level may be higher than that of a non-smoker. If so, you
should try to avoid places where there is a lot of smoke.
How can I
There are many
different ways to stop smoking, but there is no one way that's best for
everybody. The cotinine test will help you to measure the success of
whatever way you try. Ask your doctor for advice, or contact
organizations that are experienced in helping people give up cigarettes.