Biology Forum Microbiology Mitogenesis assay?

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    • #13638

      Hi, in one of my Labs we conducted a mitogenesis assay on a splenocyte sample using Concanavalin A and Lipopolysacharride as our lectins/mitogens. Different concentrations of mitogen were added to the cell suspension and the plated suspensions were measured for absorbance to measure splenocyte/lymphocyte proliferation.

      When I drew curves for the two lectins, plotting concentration against absorbance, they were roughly ( very roughly ) a bell shape. With absorbance ( cell proliferation ) rising with concentration to a certain point and then falling past that point.

      Can anyone tell me why this happens, i.e Why does cell proliferation fall after mitogen concentration rises above a certain point? I have searched the literature but have not been able to find an answer.

      I have read in a paper that mitogens/lectins can be toxic, so my current theory is that past a certain concentration the mitogen becomes toxic to the cell culture and prevents/inhibits proliferation. Am i on the right lines?? Please link papers/sources.
      Sorry for the long post, thanks.

    • #100823

      what about cell cycle? Didn’t you just observe the end of cell cycle?

    • #100893

      Yea, it turned out to be just that above a certain concentration the mitogenic effect was so pronounced that it caused the lymphocytes to proliferate to the extent that, in the time until absorbance was measured, the cells exhausted nutrients and filled the well with waste products which killed them and explained the change in absorbance.

      I was too busy searching for evidence of toxicity that i ignored the basic answer.

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