Forensic Toxicology Lab Procedures

What test is used for Amphetamines?

Marquis Reagent Test

What positive results come from a Marquis reagent test ?

orange to brown = amphetamines/Meth
dark purple/black = MDMA (Ecstasy)
purple to dark violet = Opiates (heroin, morphine, codeine)
will not react with cocaine

Marquis Test

most common test for opiates, but also for methamphetamines and some artificially produced drugs

What is the Nitric Acid Test?

2nd step after the Marquis Reagent Test to differentiate between Heroine and Morphine after receiving a purple color result

What are your positive results for the Nitric Acid Test?

Heroine = Green
Purple = Morphine

What test do you use for Marijuana/Hash/Hash Oil-THC

Duqueonis-Levine Reagent

What positive results do you receive for the Duqueonis-Levine Test?

Purple = Marijuana/Hash/Has-oil/THC

What test do you use for Oxycodone/Oxycotion/Fentanyl

Special Opiates Reagent

What positive results do you receive for the special opiates test?

violet-red = heroine
violet-red = morphine
yellow = Oxycodone
blue= codeine

What test confirms Heroine?

Mecke's Modified Reagent

What positive results do you receive for the Mecke's Modified Reagent Test

White/Brown/Black Tar = Heroine

What test do you use to test for Methamphetamine's ? When do you use it?

Methamphetamine Reagent Test

After the Marquis Reagent Test to further Methamphetamine/MDMA/Ecstasy confirmation

What positive results do you receive with the Methamphetamine Reagent Test?

Dark Blue/Dark Purple = MDMA

What does Tremodal test positive for?

Opiates

What does Wellbutrin test positive for?

Amphetamine/Methamphetamines/LSD

What does Naproxen/Ibuprofen test positive for?

Barbituates
THC
PCP

What does Percocet test positive for?

Oxycodone

What does Nifedipine test positive for?

Amphetamine

What does Amitriplyin test positive for?

LSD

What does Tizanidine test positive for?

Opiates

Scott Test

test for cocaine

Scott Test steps and results

Uses 3 different solution: 1) add 1st solution and sample turns blue 2) add solution turn pink 3) add solution= back to blue if positive for cocaine

Types of toxicology procedures?

Screening and Confirmation

What are presumptive test?

quick screening to narrow down possibilities Ex: Spot/Color test, TLC, GC, Immunoasay

What are confirmation test?

determine the exact identity Ex. chromatography, microcrystalline tests, mass spectrometry, IR Spectometry

Color Test?

Marquis Test, Scott test, Duquenois-Levine test

Types of Confirmation test?

Microcrystalline Tests, Chromatography, Mass spectromestry

Microcrystalline Test

identifies by using chemicals that react to produce characteristic crystals of specific colors and shapes (to be matched to standards)

Chromatography

separate drugs into their chemical components and used to make positive ID

•A major advantage of chromatographic techniques is that multiple xenobiotics may be detected and measured in a single procedure.
Nor is it necessary to know in advance the specific xenobiotic to be looked for. For this reason, chromatographic techniques have a major role in screening for multiple xenobiotics.

EX: TLC, HPLC, and gas

Mass Spectrometry

find chemical fingerprint. -No 2 drugs have the same chemical fingerprint because it breaks down and looks at the diff. elements in that drug/ toxin and their relative quantities -and every substance has its own elemental makeup and own molecular structure and its reflected in the chemical fingerprint

Which of the following is an alcoholic disinfectant?
Zepiran
Mercuric chloride
70% ethyl alcohol
Betadine

70% ethyl alcohol

What is the enzyme present during oxidation of gas chromatography?
NAD
NADH
Alcohol dehydrogenase
acetaldehyde

Alcohol dehydrogenase

This is a device used for measuring alcohol content of blood by analyzing alveolar breath.
Drunkometer
Intoximeter
Breathalyzer
All of the choices

All of the choices

This measures the vibrations of atoms based on the determination of functional groups.
Intoximeter
IR Spectroscopy
Gas chromatography
GC-MS Spectroscopy

IR Spectroscopy

Which of the following is used as a confirmatory test for drug/alcohol determination?
Spectrometry
Gas chromatograpy
GC-MS Spectrometry
Fuel cell devices

GC-MS Spectrometry

What is the anticoagulant used for the blood sample to be submitted in toxicology?
EDTA
Sodium fluoride
Potassium hydroxide
Lithium fluoride

EDTA

Why are spot tests are more commonly performed on urine specimens or gastric aspirates?

Because the reagents may cause precipitation of serum proteins

Why do assays suffer from fairly frequent false-positive results and occasional false-negative results?

Because of the poor selectivity of chemical reagents, as well as substantial variability in visual interpretation

Immunoassays

•The need to measure very low concentrations of an analyte with a high degree of specificity led to the development of immunoassays.

•The combination of high affinity and high selectivity makes antibodies excellent assay reagents.

What are the two common types of immunoassays?

noncompetitive and competitive

What happens in competitive immunoassays?

An analyte from the patient's specimen competes for a limited number of antibody binding sites with a labeled version of the analyte provided in the reaction mixture.

Why are drug immunoassays are usually competitive?

Most drugs are too small to have two distinct antibody binding sites

Immunoassays: EMIT

•Binding of the enzyme-labeled drug to the assay antibody BLOCKS the active site, inhibiting conversion of NADP+ to NADPH.

Free drug from the specimen can bind to the test antibody and displace it from the drug-enzyme conjugate, thereby unblocking the active site and increasing the rate of reaction.

what does EMIT stand for?

Enzyme-multiplied immunoassay technique (EMIT) test

Immunoassays: Magnetic microparticle chemiluminescent competitive assay

Magnetic beads coated with anti-drug antibody -> drug-enzyme conjugate -> substrate absorbs light-> drug in sample adding more conjugation (more drug = more competition) -> wash -> less light = less conjugation

In immunoassays the drug measured is attached to what enzyme near the active site?

enzyme glucose-6-phosphate dehydrogenase (G6PD)

different hemoglobins can be determined by

spectrochemical test

Thin Layer Chromatography:

- The stationary phase is a layer of resin on a glass or plastic plate.

- The samples are loaded at the bottom and the mobile phase migrates upward by capillary action.

- The analytes separate based on their solubility in the mobile phase.

Liquid Column Chromatography:

Stationary phase is a resin in a column. The mobile phase is a liquid solution.

Variations:
- High Performance Liquid Chromatography (HPLC)
- Fast Protein Liquid Chromatography (FPLC)
Both are performed at high pressure.

Gas Chromatography:

The mobile phase is a carrier gas, usually an inert gas such as helium or an unreactive gas such as nitrogen,

the stationary phase is a microscopic layer of liquid or polymer on an inert solid support, inside glass or metal tubing (a column).

-- Usually couple to mass spectrometry to identify the analytes.

when testing toxicity, exposure can be:

acute (single <24h)
•Subacute (multiple < 1 month)
•Subchronic (1-3 months)
•Chronic (>3 months

What are the four basic steps of the DNA extraction process?

1) Solubilization of stain components

2)Denaturation & hydrolysis of proteins

3)Removal of Denatured Proteins

4)Purification of DNA

Explain the purpose and process of each step of DNA extraction

1) Solubilization of stain components

- Add EDTA to lysis buffer; prevents Nucleases from degrading DNA

2)Denaturation & hydrolysis of proteins

- Cells are lysed using detergent SDS, Proteinase K, and DDT


3)Removal of Denatured Proteins

Denatured cells are removed using a PCIA phenol chloroform and isoamyl alcohol

4)Purification of DNA

Lysate your cells and/or tissue samples using the most appropriate procedure ex) RtPCR to isolate the nucleic acids from its contaminants into a suitable buffer solution

Qualitative Determination of Alcohol

A laboratory procedure to identify the presence of alcohol in a sample without quantifying the amount.

What is the correct procedure when preparing an acid solution?
a) Add water to acid
b) Add acid to water slowly
c) Mix acid and water at the same time
d) It doesn't matter which is added first

b

Class 3 - Flammable Liquids and Combustible Liquids

A classification of liquids that are prone to catching fire and burning easily.

Class 4 - Flammable Solids; Spontaneously Combustible Materials; Dangerous When Wet Materials

A classification of solids that are flammable, can ignite spontaneously, or react dangerously with water.

Material Safety Data Sheet (MSDS)

Contains details on chemical compounds, hazards, handling, and first aid measures.

Qualitative Tests

Tests to determine the presence and type of substances through reactions