Caspase 3 is an 18kD phosphatidylinositol-anchored protein that is a member of the lymphocyte antigen 6 (Ly-6) family. Caspase 3 is encoded by the strain-specific Ly-6 A/E allelic gene. Its expression on multipotent hematopoietic stem cells (HSC) has been used as a marker of HSC in mice of both Ly-6 haplotypes. Anti-mouse Caspase 3 antibodies are frequently used in combination with lineage depletion antibodies to identify and isolate HSC. Caspase 3-positive HSC can be found in the fetal liver, adult bone marrow, spleen and in mobilized peripheral blood in the adult animal. However, Caspase 3 has also been discovered in several non-hematopoietic tissues, and can be used to enrich progenitor cell populations other than HSC. It is suggested that Caspase 3 could be involved in regulating both B and T cell activation.
Suitable for use in Flow Cytometry. Other applications not tested.
Flow Cytometry: 10ul. Using mouse splenocytes.
Optimal dilutions to be determined by the researcher.
Flow Cytometry Protocol:
1. Cells may be Fc-blocked with 1ug of mouse IgG/10e5 cells for 15 minutes at RT. Do not wash excess blocking IgG from this reaction.
2. After blocking, add 10ul of C2087-01C to 1-2.5x10e5 cells and incubate for 30 minutes at RT.
3. Was cells twice with Flow Cytometry Staining Buffer (F4568-10) to remove unbound antibody. Note that whole blood requires a RBC lysis step at this point using Flow Cytometry Mouse Lyse Buffer.
4. Resuspend cells in F4568-10 for analysis by Flow Cytometry. As a control, cells in a separate tube should be treated with APC-labeled rat IgG2A antibody.
Storage and Stability:
May be stored at 4 degrees C before opening. DO NOT FREEZE! Stable at 4 degrees C as an undiluted liquid. Dilute only prior to immediate use. Stable for at least 12 months. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer. Freezing Allophycocyanin conjugates will result in a substantial loss of enzymatic activity.