Molecular Unit #2

Testing Considerations

type of storage and specimen collection, concerns and equipment of contamination, workflow and controls of quality control

Specimen Collection

very important, consult manufacture package, lab guidelines, CLSI

CLSI

Creates the best practices of specimen

Specimen Rejection

hemolysis, mislabeled, not collected right, and chain of custody issues

Most common specimen

blood due to nucleic acid concentration

EDTA (purple top)

ideal for DNA analysis

Heparin (green top)

limits polymerase activity

ACD (yellow top)

is good for molecular too

Specimen Types

comes from pretty much anything from the body

Contamination of Specimen

Nucleic acid from person collecting or transporting, improper storage/transport, degradation of DNA/RNA

Contamination of Equipment

amplification methods (from other samples, QC material, environment)

Good Laboratory Practices

Having physical seperation and time seperation to reduce contamination

Unidirectional Workflow

CLIA requires this, everything moves one way in a lab to prevent contamination

Open System

lab developed test, more potential for error

Closed System

everything all in one machine, reduces error. FDA approved to be done in one room

Protocol and General Practice

must be followed by everyone every single time

GLP Organization, Supplies, and Equipment

separate workspaces for each stage and dedicated equipment per station. Usually disposable equipment

Positive Pressure

used for reagent handling

Negative Pressure

used for specimen handling

Bleach and UV light

decontaminates surfaces

Quality Control

must be ran every single time a sample is ran

Target specific Assay

internal controls for each test

Multiplex Assays

control for each analyte

Qualitative Assays

positive and negative for sensitivity

PCR Based Assays

amplification controls

Reagent Preperation

first thing done in order to prevent cross contamination

Carryover Contamination

Amplified DNA/RNA from previous samples

Pre-Amplification

Reagent preparation and Specimen preparation occurs

Amplification

testing occurs

Post-Amplification

Detection of amplified product, analysis of results, specimen storage, and reporting of results occurs