SOFT 2004 Meeting Abstracts - Society of Forensic Toxicologists

M24
A MICROTITER PLATE ELISA PLATFORM FOR SCREENING ABUSE DRUGS IN ORAL FLUID
Thomas Kupiec*, Lacinda DeCicco, Susan Bernard, Nicole Vu' Analytical Research Laboratories, Oklahoma
City, OK.
The purpose of this study was to evaluate the suitability of microtiter plate ELISA assays (Neogen Corp) for
screening of abuse drugs in oral fluid (OF), and the efficiency of drug recovery from a oral sampling device,
Sarstedt Salivette®. The following drugs were tested at the proposed cutoff levels for the respective compound
according to the Federal Workplace Drug Testing Program draft guidelines: cocaine, morphine, 6­
monoacetylmorphine (6-MAM), d-amphetamine, d-methamphetamine, and methylenedioxymethamphetamine
(MDMA) and methylenedioxyamphetamine (MDA). Twenty, non-smoking, drug-free donors by testimony spit
approximately 25 mL OF into a clean cup and the sample was then poured away from the froth and centrifuged.
All OF samples were confirmed negative by GCIMS for all of the tested analytes. The samples were then pooled
to create a blank OF matrix pool used in all assays. The OF pool portions were spiked with the above analytes at
the proposed cutoff levels for the respective compound. Next, spiked or blank OF was collected from a cup
using an oral sampling device. The collector pad was held with tweezers and immersed in the cups of OF and
then processed according to the manufacturer's protocol. OF samples of 20~L were analyzed alongside a
standard curve by ELISA to determine the dilution factor needed to bring the %BlBo of each drug in OF at the
standard proposed industry cutoff levels into the optimal range for the ELISA. The calibration curve was
established by plotting log concentration VS logit transformation of the absorbance ratio for each calibrator and
the blank sample (%BlB o ). The concentrations of the analyte in the test samples were then determined from the
calibration curve based on their absorbance ratios, BlB n . The percentage of drug recovery from the oral
sampling device was calculated by comparing the concentration of the processed sample using the device to the
non-processed sample concentration. The amount of drug detected by the ELISA when no collection device was
utilized was determined by Observed/Experimental x 100. The intra-assay precision was evaluated based on the
mean absorbance ratio BlBo of the 16 replicate analyses of blank samples that were used in LOD determination.
These results are summarized in Table I. The results indicated that this ELISA platform is suitable for the
analysis of the following drugs in OF: cocaine, morphine, 6-MAM, d-amphetamine, d-methamphetamine, and
MDMA.
Table 1
I
!
I
Drug
i Dilution of
. OF
I
Drug Conc.
.
inOF
(ng/mL)
I
i
1
Amount Recovered
of OF
(ng/mL)
No Collector
Collector
Intra-assay
Precision of
BlBo
!
i Cocaine I: 50
Amphetamine I: 5 i
5
20
25
50
i
4.76±0.15
24.01±1.90
! 17.86±2.56
!
40.86±3.85
2.07±0.32
14.96±2.47
13.84±0.55
36.55±1.94
3.863
3.863
I
I MDA
No dil.
I Methamphetamine 1: 5
MDMA I: 5
Morphine I: 50
I
i 6-MAM I: 5
I
I
I
I
I
50
100
25
50
25
50
20
40
2
4
10.42±2.58
23.58±23.52
36.37±2.56
56.32±3.20
30.26±6.45
50.07±5.78
I9.62±5.71
42.36±0.96
3.13±0.77
5.67±0.72
56.24±8.96
63.19±16.74
36.30±IO.l8
58.42±2.12
10.581
2.755
22.53±7.5
44.08±6.64 I
8.316
15.18±4.43
32.79±7.43
2.43±0.19
5.56±L25
I
I
I 3.798 i
I
i
4.564
!
I
Keywords: ELISA, Oral Fluid, Drugs of Abuse
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