|28572S||1 Kit (50 assays)||
|Product Includes||Quantity (with Count)||Reactivity|
|CD3 (UCHT1) Mouse mAb (PE Conjugate) 46233||1 x 500 µl||H|
|CD4 (RPA-T4) Mouse mAb (violetFluor™ 450 Conjugate) 26755||1 x 500 µl||H|
|CD8α (RPA-T8) Mouse mAb (PE-Cy7® Conjugate) 27442||1 x 500 µl||H|
|TNF-α (D1G2) Rabbit mAb (Alexa Fluor® 647 Conjugate) 60959||1 x 100 µl||H|
|IFN-γ (D3H2) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) 12942||1 x 100 µl||H|
All reagents required for this protocol may be efficiently purchased together in our Intracellular Flow Cytometry Kit (Triton™ X-100) #51995, or individually using the catalog numbers listed below.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
NOTE: When including fluorescent cellular dyes in your experiment (including viability dyes, DNA dyes, etc.), please refer to the dye product page for the recommended protocol. Visit www.cellsignal.com for a full listing of cellular dyes validated for use in flow cytometry.
NOTE: Adherent cells or tissue should be dissociated and in single-cell suspension prior to fixation.
NOTE: Optimal centrifugation conditions will vary depending upon cell type and reagent volume. Generally, 150-300g for 1-5 minutes will be sufficient to pellet the cells.
NOTE: If using whole blood, lyse red blood cells and wash by centrifugation prior to fixation.
NOTE: Antibodies targeting CD markers or other extracellular proteins may be added prior to fixation if the epitope is disrupted by formaldehyde and/or Triton™ X-100. The antibodies will remain bound to the target of interest during the fixation and permeabilization process. Conduct a small-scale experiment if you are unsure.
NOTE: Count cells using a hemocytometer or alternative method.
posted January 2017
revised June 2020
Protocol Id: 1344
Monoclonal antibodies were purified from tissue culture supernatant via affinity chromatography. The purified antibodies were conjugated under optimal conditions, with unreacted dye removed from the preparation.
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