We are isolating CTLA-4+ cells using Miltenyi immunomagnetic beads and are finding that the cells are massively dying on the column. We don't see the other subsets isolating so believe it to be apoptosis (??) induced by CLTA-4 Ab engagement (although not what we would expect since CTLA-4 is a negative regulator of activation), and we don't see cell death after primary flow Ab staining. It's unclear if it is the column itself (but again, don't see this with other subset isolations), the primary fluorochrome-Ab staining plus the bead-anti-fluorochrome blinding or something else. We certainly need to ensure that the CTLA-4 Ab we are using is not functional or reactive.
Dear Mars
Anti-Human CD152 (CTLA-4) Biotin
Also known as: CTLA4
Clone: 14D3
RUO: For Research Use Only. Not for use in diagnostic procedures.
13-1529-82 -10 ug
Description:
The 14D3 monoclonal antibody reacts with human CD152, also known as cytotoxic T lymphocyte antigen-4 (CTLA-4). CTLA-4, a protein with structural similarities to CD28, is expressed on activated T cells (activated B cells may also express CTLA-4) and binds the B7 family members, CD80 (B7-1) and CD86 (B7-2), with higher affinity than CD28 does. CTLA-4 and CD28 appear to deliver opposing signals to T cells: while CD28 is a potent costimulator, CTLA-4 restricts the progression of T cells to an activated state by inhibiting IL-2 secretion and cellular proliferation. The cytoplasmic portion of CTLA-4 contains ER retention motifs, resulting in intracellular localization of a large proportion of newly synthesized CTLA-4 in response to TCR signaling.
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