I-O™ ALDH Activity Assay Kit, Flow Cytometry (CAT#: ITS-0622-YT382)

I-O™ Dry ALDHFLU reagent, 50 µg
I-O™ DEAB Reagent, 1.5 mM, in 95% ethanol, 1 mL
HCL, 1.5 mL
DMSO, 1.5 mL
I-O™ ALDH Assay Buffer, 4 x 25 mL

1. Assemble all necessary supplies and allow kit reagents to come to room temperature (15 - 25°C) before use.
2. Add 25 µL of DMSO to the vial of I-O™ dry ALDHFLU Reagent, mix well and let it stand for 1 minute at room temperature (15 - 25°C).
3. Add 25 µL of 2 N HCl and mix well. Incubate this mixture for 15 minutes at room temperature (15 - 25°C).
4. Add 360 µL of I-O™ ALDH Assay Buffer to the vial and mix.
5. Keep the activated ALDHFLU Reagent at 2 - 8°C during use.
6. Aliquot the remaining activated ALDEFLUOR™ Reagent and store at -20°C.

1. Prepare a "test" and a "control" tube for each sample to be tested. Place 1.0 mL of the cell suspension into each "test" sample tube.
2. Add 5 µL of I-O™ DEAB Reagent to the "control" tube. Recap control tube and DEAB vial immediately.
3. Add 5 µL of the activated I-O™ ALDHFLU reagent per milliliter of sample to the first sample "test" tube. Mix and immediately transfer
0.5 mL of the mixture to the DEAB "control" tube.
4. Add control and substrate solutions as described in steps 2 and 3 above for each sample to be tested.
5. Incubate samples for 30 to 60 minutes at 37°C (do not exceed 60 minutes).
6. Centrifuge all tubes for 5 minutes at 250 x g and remove supernatant. Resuspend cell pellets in 0.5 mL of
I-O™ ALDH Assay Buffer and store the cells on ice or at 2 - 8°C.
7. Optional: Perform a viability cell count or immunophenotyping.
8. Flow cytometry analysis.

Mammals, Other species

For unused kit: All the reagents should be kept at 2 - 8°C.
For used kit: Aliquot the activated ALDEFLUOR™ Reagent and store at -20°C.

Stable as supplied until expiry date on label.

Restrictions for research use only. Intended for use by laboratory professionals.