Laboratory Techniques & Safety Flashcards

A blood-derived DNA sample repeatedly fails to amplify while controls work. What is a quick first step to test for inhibition?

Answer: Dilute the template and re-run the reaction

Diluting the template reduces inhibitor concentration (e.g., heme, heparin) and often restores amplification.

What is the primary purpose of an internal amplification control spiked into each sample?

Answer: To distinguish a true negative result from a failed reaction due to inhibition

If the internal control amplifies but the target does not, the negative result is valid rather than caused by inhibition.

Which of the following substances is a well-known PCR inhibitor commonly carried over from blood collection tubes?

Answer: Heparin

Heparin binds polymerase and DNA, inhibiting amplification; MgCl2 is a cofactor and BSA/trehalose are enhancers.

Why is a manual-defrost (non-frost-free) freezer preferred for storing PCR enzymes and primers?

Answer: Frost-free freezers cycle temperature, causing repeated partial thawing of reagents

Defrost cycles in frost-free units warm contents periodically, degrading enzymes and oligonucleotides.

What is the main benefit of aliquoting primer and dNTP stocks into single-use volumes?

Answer: It limits freeze-thaw cycles and confines any contamination to a single aliquot

Small aliquots preserve reagent integrity and make contaminated stock easy to discard without losing everything.

A tube of patient specimen breaks inside a centrifuge. What is the correct immediate action?

Answer: Keep the lid closed for about 30 minutes to let aerosols settle, then decontaminate with appropriate PPE

Waiting allows aerosols to settle before opening, followed by careful disinfection per biosafety procedures.

What is a key hazard when using liquid nitrogen or large amounts of dry ice in a poorly ventilated room?

Answer: Oxygen displacement leading to asphyxiation

Evaporating nitrogen or sublimating CO2 can displace oxygen in confined spaces.