Step-1 |
- Dispense 20 μl denaturation (DEN) solution in the corner of each trough.
Note: No need to change tips between samples.
|
Step-2 |
- Add 20 µl Amplicon to DEN and carefully mix by pipetting up and down.
- Incubate for 5 minutes.
Note: Change tips between samples.
|
Step-3 |
- Carefully add 1 ml of pre-warmed hybridization (HYB) buffer to each well, using filtered tips.
Note: Change tips between samples.
- Gently shake the tray to homogenously mix HYB with amplicon.
- Do not spill into neighbouring troughs.
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Step-4 |
- Carefully place numbered strips in each well, strip face side up using clean/ new plastic tweezers.
- Strips must be completely immersed in HYB.
- If strips turn over, reposition them with a fresh pipette tip.
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Step-5 |
- Place tray onto Twincubator and press START.
- Twincubator will then start shaking with a temperature of 45°C.
- This step will be of 30 minutes.
- After 30 minutes, the Twincubator will set off an alarm.
- When the alarm goes off, press the right arrow to stop.
|
Step-6 |
- Completely aspirate HYB using transfer pipettes.
Note:
Change pipettes for every sample.
Do not decant the contents of the tray into a sink or jar as this can contaminate the work area.
Wipe off the condensation that forms on the lid of the incubator before every incubation step.
|
Step-7 |
- Add 1 ml of STR solution to each well.
- Press start.
- This step will be of 15 minutes at 45°C.
- After 15 minutes, the Twincubator will set off an alarm.
- When the alarm goes off, press the right arrow to stop
|
Step-8 |
- Completely aspirate STR solution by transfer pipette.
Note:
Change pipettes for every sample.
Do not decant the contents of the tray into a sink or jar; this can contaminate the work area.
|
Step-9 |
- Add 1ml of RIN solution to each well.
- Press start.
- This step will be for 1 minute.
- After 1 minute, the machine will set off an alarm.
- Once the alarm goes off, press the right arrow to stop.
|
Step-10 |
- Completely aspirate the RIN solution by transfer pipette.
Note: No need to change pipette between wells.
- From this step, the content of the tray may be decanted into a container with 1% sodium hypochlorite solution.
|
Step-11 |
- Add 1 ml of conjugate solution to each well using a micropipette.
Note: No need to change tips for every sample.
- Press start.
- This step will be of 30 minutes.
- After 30 minutes, the machine will set off an alarm.
- Once the alarm goes off, press the right arrow to stop.
|
Step-12 |
- Completely aspirate conjugate solution using a transfer pipette, or the content of the tray may be decanted into a container with 1% freshly prepared sodium hypochlorite solution.
|
Step-13 |
- Work at room temperature from this step.
- Add 1 ml of RIN solution to each well.
- Press start.
- This step will be for 1 minute.
- After 1 minute, the machine will set off an alarm.
- Once the alarm goes off, press the right arrow to stop.
|
Step-14 |
- Completely aspirate the RIN solution using a transfer pipette, or the content of the tray may be decanted into a container with 1% freshly prepared sodium hypochlorite solution.
|
Step-15 |
|
Step-16 |
|
Step-17 |
- Add 1 ml of distilled water to each well.
- Press start.
- This step will be for 1 minute.
- After 1 minute, the machine will set off an alarm.
- Once the alarm goes off, press the right arrow to stop.
|
Step-18 |
- Completely aspirate distilled water using a transfer pipette, or the content of the tray may be decanted into a container with 1% freshly prepared sodium hypochlorite solution.
|
Step-19 |
- Add 1 ml of substrate solution to each well.
Note: No need to change tip.
- Cover the machine with aluminium foil as the substrate solution is photosensitive.
- Press start.
- This step may be for 3-5 minutes (depending on the colour that develops).
- The machine will set off an alarm.
- Once the alarm goes off, press the right arrow to stop.
|
Step-20 |
- Completely aspirate the substrate solution using a transfer pipette, or the content of the tray may be decanted into a container with 1% freshly prepared sodium hypochlorite solution.
|
Step-21 |
- Add 1 ml of distilled water to each well.
- Press start.
- This step will be for 1 minute.
- After 1 minute, the machine will set off an alarm.
- Once the alarm goes off, press the right arrow to stop.
|
Step-22 |
- Completely aspirate distilled water using a transfer pipette, or the content of the tray may be decanted into a container with 1% freshly prepared sodium hypochlorite solution.
|
Step-23 |
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Step-24 |
- Do not aspirate distilled water from wells.
|
Step-25 |
- Using clean tweezers, remove the strips from the tray and dry them between two layers of absorbent paper
|
Step-26 |
- Align strips and paste them on a reporting sheet using cello tape/ scotch tape.
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Comments
Steps can be represented…
Dr. Priyadarshini Fri, 12/01/2024 - 06:51
Steps can be represented better - flowchart or video.