If antibodies are used for cell sorting, how long do they remain on the cell? How long does it take for these surface markers to be turned over? I am unsure if once cells are sorted if one can eventually assume the antibodies are no longer on the cells. I have been unable to find much information on antibody recycling/removal rate, and I am wondering if there is existing literature on this.
Hi Olivia.
It is a wild question to answer in a specific manner.
It may depends on many factors including i) the use of fixed or live cells for antibody labelling, ii) the cell phenotypes used, i.e., active cells in the activity of endomembrane system (largely, highly secreting cells with exo- and endocytosis) at active state, iii) the properties of target molecules labelled, i.e., whether the antibody binding causes endocytosis and subsequent vesicles to be fused with lysosomes for degradation, and iv) the distribution of cytoskeletal (actins) and other specific proteins underlying the plasma membranes.
Thus, no such a generalized report is available. If the cells are active in secretion, a type II transmembrane protein that I studied in CHO cells and neurons, the labelled antibodies to the molecules found in many different levels of cyctoplamic endomembrane system, beginning to show Golgi accumulation of the labelled proteins in live cells at 15 min post-labelling forwards under culture condition.
It can be stopped by placing dish at 4C, and cytochalasin D treatment at 37C.
Therefore, it may be correct to say recycling molecules on the plasma membranes in endomembrane system, but not recycling of the antibody labelled. Labelled antiboy molecules may just be degraded, but not recycled in less-active cells and low temperature for labelling.
Relabelling cells with another marker may not just be a great idea.
Best wishes.
If you keep everything (including the sorter) at 4 degrees C you will not have any appreciable turnover of cell surface receptors. At room temp or warmer it depends on the receptor and the antibody used. You might get away with a Fab since it will not crosslink.
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