TBMNK Flow Cytometry Assay Kit

The RayBio^® Human TBMNK Flow Cytometry Kit is designed to determine the percentages of mature human lymphocyte and monocyte subsets in peripheral whole blood (WB). The lymphocyte subsets include T lymphocytes (CD3⁺), Helper T lymphocytes (CD3⁺CD4⁺), Cytotoxic T lymphocytes (CD3⁺CD8⁺), B lymphocytes (CD19⁺) and a subsets of Natural killer (NK) lymphocytes (identified by CD56 and CD16). The monocyte subsets include classical monocytes (CD14^brightCD16^-), intermediate monocytes (CD14^brightCD16⁺), and non-classical monocytes (CD14^dimCD16⁺). Our kit contains individually packed fluorochrome-labeled antibodies, antibody diluent, and lysing solution to minimize the red blood cells. This kit is easy and convenient to use, and a perfect tool for immunophenotyping peripheral lymphocytes and monocytes of human whole blood.

• RayBright^® Violet 450 Anti-human CD19
• RayBright^® Violet 500 Anti-human CD4
• RayBright^® Blue 488 Anti-human CD3
• PE Anti-human CD14
• PerCP Anti-human CD16
• APC Anti-human CD56
• RayBright^® Red 700 Anti-human CD45
• RayBright^® Red 780 Anti-human CD8
• Antibody Diluent
• RayBio^® 10X RBCs Lysing Buffer

• Flow cytometer with violet, blue and red lasers
• 1.5 mL polypropylene microcentrifuge tubes or similar
• 12 × 75-mm tubes
• De-ionized (DI) H₂O
• Centrifuge
• Vortex mixer

1. Dilute the RayBio^® 10X Whole Blood Lysing Solution 10-fold with de-ionized (DI) H₂O to prepare 1X lysing solution.
2. Prepare antibody cocktail in a 1.5 mL tube by adding 2.5 μL of the antibody reagents one by one and 30 μL of the Antibody Diluent in the kit per single test. The final volume of antibody cocktail per single test is 50 μL.
3. Label each 12 × 75-mm tube and add 50 μL of well-mixed, anticoagulated whole blood from each sample to the bottom of the tube.
4. Pipette 50 μL of antibody cocktail from step 2 above into the bottom of each tube and vortex gently to mix.
5. Incubate for 20 minutes in the dark at room temperature (20°C–25°C).
6. Add 1 ml of 1X lysing solution from step 1 to each tube and vortex gently to mix.
7. Incubate for 20 minutes in the dark at room temperature (20°C-25°C).
8. Centrifuge the samples at 500 x g for 5 minutes and discard the supernatant without disturbing the pellet.
9. Add 0.5 mL of 1X lysing solution from step 1 to each tube. The sample is now ready to be analyzed on a flow cytometer.

Store kit at 4°C, protected from light for up to three months for full functionality.