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Hybridization Setup in LPA and Genotypic DST
Learning ObjectivesHybridization Setup in LPA and Genotypic DST
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The steps listed below need to be followed when setting up for the hybridisation procedure.
Workstation Set-up
- Compile all the tools and reagents required for the step.
- Switch on all the equipment and check whether they are in working condition.
- Always connect the Twincubator and GT blot to the UPS.
- Thoroughly clean the workbench and tools with appropriate disinfectant.
- Clean the equipment as suggested by the manufacturer.
- Place an absorbent sheet.
- Bring out the hybridization reagents from the refrigerator.
Reagent Preparation
- Kit Component 1 is used for this step.
- Hybridization reagent preparation and steps involved in hybridization procedure remains the same in First Line (FL) and Second Line (SL) Line Probe Assay (LPA).
- Bring out the reagent kit from the refrigerator.
- Pre-warm the hybridisation and stringent solution in a water bath, as these 2 solutions form crystals due to the kit’s storage at 2 - 8ºC.
Reagents Required for Hybridization
- Hybridization Solution - HYB
- Stringent Solution - STR
- Conjugate Solution - CON
- Substrate Solution - SUB
- RIN Solution - RIN
- Distilled Water - DW
The steps for preparing these reagents are shown in the tables below.
PREPARATION OF HYBRIDIZATION SOLUTION | PREPARATION OF STRINGENT SOLUTION |
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PREPARATION OF CONJUGATE SOLUTION | PREPARATION OF SUBSTRATE SOLUTION |
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PREPARATION OF RIN SOLUTION | FOR DISTILLED WATER |
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Please click the video below to see more on hybridization.
Resources
- GenoType MTBDR plus ver 2.0 Kit, Instructions for Use.
- GenoType MTBDRsl VER 2.0 Kit, Instructions for Use.
- GLI Training package on LPA.
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Comments
Table 2 can be moved to next…
Dr. Priyadarshini Fri, 12/01/2024 - 06:42
Table 2 can be moved to next slide