I've activated mouse T cells with 4.5 um latex bead from polysciences. Just coat them with anti-CD3/28 and block with BSA. Try different ratios, starting at 1:1-1:10 beads: T cells

These two types of particles use different mechanisms of coupling of ligands. These differences may have special impact on orientation of bound antibodies.

Tosylactivated reacts with ligands containing primary amino or sulphydryl groups.

Epoxy - with amino, thiol and hydroxyl functional groups. (So, there are some restriction in using buffers containing Tris, sugars, etc.)

Thanks for your answers.

@Dmitry: Do you know which one is more suitable for the activation of T cells?

@Burcu: I intend to compare different costimulatory molecules to activate T lymphocytes. So a general protocol for antibody coating would be useful.

Dear Leonardo, sorry! I have not direct experience in this field. In my history, I used different particles and chemical mechanisms to prepare immunosorbents for purification of proteins. Now, I do not see any reasons to prefer one or another. I think, in both cases you will be successful in conjugation of mAbs with particles. Both are not optimal, because they contain too high density of reactogenic groups, which need blocking after coupling of antibodies. I think, the most serious problem will be to find optimal (sufficiently low) density of mAbs, resulting to appropriate activation of T cells. This will be time-consuming. My advice to you is to get collaboration with the person, who has passed this way recently.

While I have no direct answer for your specific question, I would warn you that all T cells are not equal in their responses and all antibodies are not equal either!

J Exp Med. 1984 Aug 1;160(2):494-505.

T cell antiidiotypic antibodies reveal differences between two human leukemias.

Posnett DN, Bigler RD, Bushkin Y, Fisher DE, Wang CY, Mayer LF, Chiorazzi N, Kunkel HG.

@Joseph : How did you block with BSA? Did you mix the antibodies with BSA and add to cells? Also what BSA concentration were you often using for blocking anti-CD3 or anti-CD28? Many thanks!

Dynabeads coated with aCD3/aCD28 can be tried alon with the other two beads for your experiments and tell us which is the best . Lalitha Kabilan

Hi,

In this product both clone and the ratio of anti-CD3 and anti-CD28 have been optimized for providing more physiological activation of T-cells.

kind regards

Ketil