FITC label procedure for Toxoid???
ksabelko at PHARMDEC.WUSTL.EDU
Mon Apr 24 19:51:37 EST 1995
pemanuel at umabnet.ab.edu (Peter) wrote:
> I have a toxoid proetin which I must label with FITC. I am seeking a
>procedure which I can use to purify the unbound away from the labelled
>stuff. I have been given a procedure which involves a lot of columns. It
>is also dated 1976. I cannot believe that no one has developed a better
>way than running a set of columns.
> Does anyone know of a good procedure paper on FITC conjugation?
>Any references or advice from experienced FITC users would be
>appreciated. No comment is too trivial for me. Well almost no comment.
>Anyway let me know if there is any easier way I can look into.
To purify the FITC conjugated material you might want to try dialysis.
Current Protocols in Immunology recommends dialyzing in 500 ml dialysis
buffer (0.1M Tris.Cl, pH7.4, 0.1% (v/v) NaN3, 0.2 M NaCl; pH adjusted to 7.4
with NaOH) @ 4 C x 48 hrs, with 2-3 changes of buffer. You can easily
monitor the dialysis by the change in color of the buffer -- the first 1-2
changes it will be yellow.
Reference: Current Protocols in Immunology, Chp. 5 Immunofluorescence and
Cell Sorting, pp 5.3.2-5.3.3
Good luck-- Kim
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