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Drugs of abuse: Methamphetamine Direct (MDMA, Ecstacy),ELISA kit
/Product Detail : BQ 090D Drugs of abuse: Methamphetamine Direct (MDMA, Ecstacy),ELISA kit
Product name : Drugs of abuse: Methamphetamine Direct (MDMA, Ecstacy),ELISA kit
Catalog number : BQ 090D
Quantity: 96 tests
Supplier name : Bioquant
Data sheet : Ask more or other datasheet now !
INTENED TO USE
The Methamphetamine Direct ELISA Kit 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/ mass spectrometry (GS-MS) is the preferred confirmatory method. Professional judgment should be applied to any drug of abuse test result, particularly when preliminary positive results are used.
SUMMARY AND EXPLANATION
The Methamphetamine Direct ELISA Kit is a specific and sensitive in-vitro test to detect the presence of d-methamphetamine in samples such as whole blood, oral fluids, serum, plasma and urine. While the assay will detect amphetamine use, interference by l-methamphetamine and pseudo-ephedrine is virtually nonexistent. Methamphetamine is a potent central nervous system stimulant with less peripheral actions than mphetamine. The (+)-isomer also referred to as d-methamphetamine is ten times more potent than the (-)-isomer, l-methamphetamine. Amphetamines act by inducing euphoria, irritability, anxiety and paranoia Methamphetamine is metabolized to its active metabolite amphetamine (via N-demethylation) and is further metabolized by hydroxylation and deamination of amphetamine. Urinary excretion rates are influenced by the urinary pH with acidic urine favoring the excretion of unchanged drug. Alkaline urine reduces the excretion of unchanged methamphetamine to less than 5% of the dose.
PRINCIPLES OF THE TEST
The Methamphetamine Direct ELISA Kit (for d-methamphetamine measurement) is based upon the competitive binding to antibody of enzyme labeled antigen and unlabeled antigen, in proportion to their concentration in the reaction mixture. A 10 ěl. aliquot of a diluted unknown specimen is incubated with a 100 ěl. dilution of enzyme (Horseradish peroxidase) labeled d-methamphetamine derivative in micro-plate wells, coated with fixed amounts of oriented high affinity purified polyclonal antibody. The wells are washed thoroughly and a chromogenic substrate added. The color produced is stopped using a dilute acid stop solution and the wells read at 450 nm. The intensity of the color developed is inversely proportional to the concentration of drug in the sample. The technique is sensitive to 1 ng/ml. The Methamphetamine Direct ELISA Kit avoids extraction of urine sample for measurement. It employs a d-methamphetamine directed antiserum. Due to the proprietary method of orienting the antibody on the polystyrene micro-plate much higher sensitivity is achieved compared to passive adsorption. This allows an extremely small sample size, reducing matrix effects and interference with binding proteins(s) or other macromolecules.
MATERIALS NOT PROVIDED
1. 12x75 mm Disposable Glass or Plastic Culture Tubes to pre-dilute samples (if required).
2. Manual or electronic micropipettes (single channel or multi channel) or automated pipetting stations.
3. Refrigerator (for kit storage).
4. Interval Timer.
5. Wash bottle or Plate Washer.
6. Micro-plate reader capable of reading at 450 nm and 650 nm.
STORAGE AND STABILITY
1. Urine samples should be stored at 2 - 4 degrees centigrade until use. Samples should be well mixed before assay. Repeated freezing and thawing should be avoided. Urine samples should be shipped refrigerated with Blue Ice or equivalent.
2. The expiration date of the kit is stated on the label.
3. The kit can be expected to perform satisfactorily until the expiration date if stored in the refrigerator at 2 – 40 C.
WARNINGS AND PRECAUTIONS
1. Potential biohazardous materials:
The calibrator and controls contain human source components which have been tested and found non-reactive for hepatitis B surface antigen as well as HIV antibody with FDA licensed reagents. However, there is no test method that can offer complete assurance that HIV, Hepatitis B virus or other infectious agents are absent. These reagents should be handled at the Biosafety Level 2, as recommended in the Centers for Disease Control/National Institutes of Health manual, "Biosafety in Microbiological and Biomedical Laboratories" 1984.
2. This kit is designed for Research Use Only.
3. Optimal results will be obtained by strict adherence to the test protocol. Precise pipetting as well as following the exact time and temperature requirements is essential.
4. Do not pipette by mouth. Do not smoke, eat, or drink in the areas in which specimens or kit reagents are handled.
5. The components in this kit are intended for use as an integral unit. The components of different lots should not be mixed.
6. Control sera and sample diluent contain preserved with sodium azide. Sodium azide may react with lead and copper plumbing to form explosive metal azide. On disposal, flush with a large volume of water.
SPECIMEN COLLECTION HANDLING
1. The Methamphetamine Direct ELISA Kit is to be used with human samples, such as whole blood, oral fluids, serum, urine and plasma. Has not tested all possible applications of this assay. Cutoff criteria are important in deciding the sample dilution.
2. Specimens to which sodium azide has been added affect the assay.
All reagents must be brought to room temperature (18-260 C) before use.
The procedure as described below may be followed in sequence using manual pipettes. Alternatively all reagents may be added using an automated pipettor.
1. Dilute specimens, to the necessary range with Phosphate Buffer Saline pH 7.0. (Urine samples are normally diluted 1:20 for a methamphetamine cutoff of 500 ng/ml.) The dilution factor and volume added can be adjusted based on the laboratory’s cutoff.
2. Add 10 ěl. of appropriately diluted calibrators and standards to each well in duplicate.
3. Add 10 ěl. of the diluted specimens in duplicate (recommended) to each well.
4. Add 100 ěl of the Enzyme Conjugate to each well. Tap the sides of the plate holder to ensure proper mixing.
5. Incubate for 60 minutes at room temperature (18-260 C) preferably in the dark, after addition of enzyme conjugate to the last well.
6. Wash the wells 6 times with 350 ěl. distilled water using either a suitable plate washer or wash bottle taking care not to cross contaminate wells. If testing samples containing abnormally high amounts of hemoglobin (some Postmortem samples), use 10 mM Phosphate buffered saline pH 7.0-7.4. This will lower potential nonspecific binding of hemoglobin to the well, thus lowering background color.
7. Invert wells and vigorously slap dry on absorbent paper to ensure all residual moisture is removed. This step is critical to ensure that residual enzyme conjugate, does not skew results. If using an automated system, ensure that the final aspiration on the wash cycle aspirates from either side of the well.
8. Add 100 ěl. of Substrate reagent to each well and tap sides of plate holder to ensure proper mixing.
9. Incubate for 30 minutes at room temperature, preferably in the dark.
10. Add 100 ěl. of Stop Solution to each well, to change the blue color to yellow.
11. Measure the absorbance at a dual wavelength of 450 nm and 650 nm.
12. Wells should be read within 1 hour of yellow color development.
13. The following data represent a typical dose/response curve.
The dose/response curve shown above should not be used in assay calculations. It is recommended that at least one in-house positive quality control sample be included with every assay run. A dose response curve or a cutoff calibrator should be run with every plate.
If the average sample absorbance is equal to or less than the average absorbance of the laboratory positive reference standard the sample is POSITIVE for methamphetamine. If the average sample absorbance is greater than the average absorbance of the laboratory positive reference standard the sample is called NEGATIVE for methamphetamine. Alternatively a dose response curve can be established by plotting standard concentration (abscissa) against corresponding absorbance (ordinate). Values for unknown samples are obtained by interpolation from the curve.
Sixty whole blood samples and 40 urine samples collected from presumed non-users were tested in the Methamphetamine Direct ELISA Kit. One hundred percent of these normal samples measured negative at 50 ng/ml for whole blood and 500 ng/ml for urine. Fifty five whole blood samples which were previously confirmed positive for methamphetamine by GC-MS employing a cut-off of 50 ng/ml, were tested in the ethamphetamine Direct ELISA Kit . All of the samples were found to be positive i.e. above the cut-off of 50 ng/ml.
The precision of the Methamphetamine Direct ELISA Kit has been verified by assessment of the mean, standard deviation (SD) and coefficients of variation (CV) in data resulting from repetitive assays.
3. Intra-assay Precision
Intra-assay precision was determined with reference controls.
A 0,10, 25 and 50 ng/ml standard was assayed five times in the same assay.
Assay sensitivity based on the minimum methamphetamine concentration required to produce a four standard deviation from assay Ao is 1 ng/ml.
The specificity of the ELISA Methamphetamine was determined by generating inhibition curves for each of the compounds listed below The antisera cross-reactivities are listed in Table .
6. Cross-Reactivity with Unrelated Drugs
Aliquots of a human urine matrix were spiked with the following compounds at a concentration of 10,000 ng/ml. None of these compounds gave values in the assay that were equal to or greater than the assay sensitivity level (1 ng/ml).
Acetaminophen, Acetylsalicylic acid, Aminopyrine, Ampicillin Amobarbital, Ascorbic acid, Atropine, Barbital, Benzoylecgonine, Butabarbital, Caffeine , Cocaine, Carbamazepine, Codeine, Chloroquine, Chloropromazine, Carbromal, Desipramine, Dextromethorphan, Dextropropoxyphene, 5,5-Diphenylhydantoin, 10-11-Dihydro carbamazepine, Diazepam, Ethosuximide, Estriol, Estrone, Estradiol, Ethotoin, Glutethimide, Hexobarbital, Ibuprofen, Imipramine, Lidocaine, LSD, Methadone, Methadone-primary metabolite, Methaqualone, Metharbital , Mephenytoin, "-Methyl-"-propylsuccinimide, Mephobarbital, Methyl PEMA, Methsuximide , 4-Methylprimidone, Morphine, Meperidine, Niacinamide, Norethindrone, N-Normethsuximide, Phenobarbital, Phensuximide,PEMA, Primidone, Phencyclidine, Pentobarbital, Phenothiazine, Procaine, Quinine, Secobarbital, Tetracycline, Tetrahydrozoline, THCCOOH.
1. Urine Testing for Drugs of Abuse, National Institute on Drug Abuse Research Monograph, 73,1986.
2. R.C. Baselt. In : Advances in Analytical Technology, Vol.1. Randall C. Baselt edd. (Biomedical Publications, Foster City, CA. 87-93).
3. Driscoll, R.C., Barr, F.S., Gragg, B.J. and G.W. Moore. Determination of Therapeutic Blood Levels of Methamphetamine by GC. J.Pharm. Sci 60:1492.1971.
All of Bio-Quant ELISA Kits have not been tested for clinical use and are not approved in the United States by the FDA for diagnostic clinical use. They are components or reagents made solely for research use, further manufacturing and export use. It is the commitment of Bio-Quant customers to receive its products solely for the purpose of exportation or research, and not for the purposes of clinical diagnostic use.
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