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Evaluation of First-line Line Probe Assay Strip
Learning Objectives- Describe the Key Zones of the FL - LPA Evaluation Strip.
- Elaborate key considerations during the evaluation of an LPA strip.
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Each strip for First Line – Line Probe Assay (FL - LPA) has a total of 27 reaction zones (Figure below).
The Key Zones Include:
Conjugate Control (CC)
- Indicates the efficiency of conjugate binding and substrate reaction
Amplification Control (AC)
- Develops when control amplicon binds indication correct procedure was followed
- The absence of AC zone indicates an invalid test
M. tuberculosis Complex (TUB)
- Hybridizes with amplicons generated from Mycobacterium tuberculosis complex
- The absence of TUB zone indicates the absence of M. tuberculosis complex
Locus Controls (rpoB, katG, and inhA)
- Detect gene region specific for genes rpoB/ katg/ inhA
Wild-type Probes
- Comprise the resistance regions of respective genes (rpoB/ katg/ inhA)
- The absence of signal for at least one of the wild type probes indicates the resistance of the tested strain to the respective antibiotic
Mutation Probes
- Detect most common resistance-mediating mutations (rpoB/ katg/ inhA)
- Each pattern deviating from the wild-type pattern indicates resistance
Figure: Key Zones of FL - LPA Evaluation Strip; Source: GenoType MTBDR plus ver 2.0 kit, Instructions for Use.
Evaluation of Assay Strip for FL - LPA Zones:
- The resistance status is determined and noted for each respective column.
- Distances between single probes on the strips may vary slightly.
- Not all bands of a strip have the same signal strength.
- The DNA strip should only be interpreted when:
- AC, CC and TUB bands are present
- Along with the presence of locus control bands for rpoB, katG and inhA
- Only those bands whose intensities are about as strong as, or stronger than that of the AC zone are to be considered.
- Bands whose intensities are about as strong as or stronger than that of the AC zone are considered.
Important: For an accurate evaluation, align provided template separately for each locus with the respective Locus Control band.
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