ASCP MB flashcards

32 cards. Read the term and the definition here. The deck at the top of this page is optional.

Central dogma

DNA is transcribed into RNA, which is translated into protein; reverse transcriptase can synthesize complementary DNA from an RNA template.

Chargaff's rule

In double-stranded DNA, adenine equals thymine and guanine equals cytosine; base pairing is A-T with two hydrogen bonds and G-C with three.

Primer Tm estimate

For short primers, Tm approximates 2°C per A or T plus 4°C per G or C; nearest-neighbor calculations are more accurate for longer oligonucleotides.

Copy number formula

Copies per microliter equal DNA mass in nanograms times 6.022×10^23 divided by fragment length in base pairs times 660 times 10^9.

Gel electrophoresis direction

Negatively charged DNA migrates toward the positively charged anode; smaller fragments move faster through the gel matrix than larger fragments.

Sanger sequencing method

Dideoxynucleotides terminate chain elongation; fluorescently labeled fragments are separated by capillary electrophoresis and detected to determine the DNA sequence.

EDTA tube use

Lavender-top EDTA tubes chelate calcium and inhibit coagulation; EDTA plasma or whole blood is preferred for many molecular assays because heparin can inhibit amplification.

Heparin interference

Heparin can inhibit polymerase chain reaction and other enzymatic steps; heparinized specimens should be avoided or treated when molecular testing is required.

FFPE nucleic acid quality

Formalin fixation crosslinks proteins and fragments nucleic acids; FFPE DNA often shows low yield and short fragments, requiring robust extraction and small amplicon assays.

Silica extraction principle

In chaotropic salt and alcohol, nucleic acids bind silica membranes; contaminants are washed away and purified nucleic acid is eluted in low-salt buffer or water.

A260/A280 ratio

Pure double-stranded DNA typically has an A260/A280 ratio near 1.8, while pure RNA is near 2.0; lower ratios suggest protein or phenol contamination.

PCR exponential phase

During the exponential phase, product doubles each cycle and amplification efficiency is maximal; quantification is most reliable before reagents become limiting.

Hot-start PCR

Hot-start enzymes are inactive at low temperatures, preventing primer extension during setup; this reduces nonspecific products and primer dimers before the initial denaturation step.

TaqMan probe mechanism

A hydrolysis probe carries a reporter and quencher; polymerase exonuclease activity cleaves the probe during extension, separating reporter from quencher and generating fluorescence.

SYBR Green detection

SYBR Green fluoresces when bound to double-stranded DNA; it detects any amplified product, so melt-curve analysis is needed to distinguish specific products from primer dimers.

qPCR efficiency formula

Amplification efficiency equals 10 raised to negative one divided by the standard-curve slope, minus one; a slope of −3.32 corresponds to 100% efficiency.

Hybridization stringency

Increasing temperature or decreasing salt concentration raises stringency, favoring perfectly matched probe-target hybrids and reducing signal from mismatched sequences.

NAT window period

Nucleic acid testing detects infection earlier than antibody-based tests because pathogen nucleic acid appears before a detectable immune response, shortening the diagnostic window period.

16S rRNA gene testing

Broad-range bacterial detection often targets conserved 16S ribosomal RNA gene regions flanking variable sequences; sequencing identifies bacteria but does not by itself prove viability.

Viral load reporting

Viral load is reported as nucleic acid copies or international units per milliliter, often as log10 values; trends over time guide treatment response.

HIV drug resistance

HIV resistance testing sequences reverse transcriptase, protease, and integrase regions to detect mutations associated with reduced susceptibility to antiretroviral drugs.

False-positive NAT causes

Amplicon contamination, cross-contamination between specimens, and carryover from positive controls can cause false-positive nucleic acid amplification results; unidirectional workflow reduces risk.

HGVS variant nomenclature

HGVS nomenclature describes variants by reference sequence and position, such as c. for coding DNA and p. for protein, enabling unambiguous reporting of genetic changes.

Variant allele fraction

Variant allele fraction equals variant reads divided by total reads at that locus; germline heterozygous variants are usually near 50%, while somatic variants often show lower fractions.

ACMG variant classes

Clinical variant classification uses pathogenic, likely pathogenic, variant of uncertain significance, likely benign, and benign categories, based on evidence such as population frequency and functional data.

BCR-ABL1 monitoring

The Philadelphia chromosome creates BCR-ABL1; quantitative reverse-transcription PCR measures transcript levels and tracks response to tyrosine kinase inhibitor therapy in chronic myeloid leukemia.

NIPT cell-free DNA

Maternal plasma contains cell-free DNA from the placenta and mother; sequencing can estimate fetal trisomy risk for chromosomes 21, 18, and 13 without invasive sampling.

No-template control

A no-template control contains all reagents except nucleic acid; amplification in this control indicates contamination and invalidates the run's negative results.

Limit of detection

The limit of detection is the lowest analyte concentration detected in at least 95% of replicate tests; it is established during validation with multiple replicates.

Unidirectional workflow

Molecular laboratories separate reagent preparation, specimen processing, amplification, and detection into distinct areas with unidirectional movement to prevent amplicon contamination.

Proficiency testing

Proficiency testing uses blinded external samples to assess laboratory performance; unacceptable results require investigation, corrective action, and documentation.

BSL-2 precautions

Most clinical molecular work uses BSL-2 practices, including gloves, lab coats, eye protection, biosafety cabinets for aerosols, and disinfection of work surfaces.