[lysine]

Hello everybody,

I would like to know more about the TNBS Assay that is used to assess the amount of free lysines in a protein, like in this protocol:

http://cmdr.ubc.ca/bobh/methods/ESTIMATIONOFAMINOGROUPSUSINGTNBS.html

1) Is it measuring the total amount of lysines or only the ones accessible (on the surface)?

2) What material would you use for the reaction tubes? I heard everything from "use only glass" to "use PS plastic tubes" or the standard PP tubes.

Thanks
lysine

[Babcock_Hall]

Gary Means and Robert Feeney (Chemical Modification of Proteins, Holden Day, 1971) have a nice discussion of this reagent in two places in their book.  It seems as if there can be unusually fast- and slow-reacting lysines.  You might also wish to consult this article, which cites Fields' work:  "The determination of ϵ-amino groups in soluble and poorly soluble proteinaceous materials by a spectrophotometrie method using trinitrobenzenesulfonic acid"  Analytical Biochemistry, Volume 207, Issue 1, 15 November 1992, Pages 129–133  http://www.sciencedirect.com/science/article/pii/0003269792905137

See also:  "The Quantification of Protein Amino Groups by the Trinitrobenzenesulfonic Acid Method: A Reexamination"  http://dx.doi.org/10.1006/abio.1997.2161,

And "Quantitation of ϵ-amino group using amino acids as reference standards by trinitrobenzene sulfonic acid: A simple spectrophotometric method for the estimation of hapten to carrier protein ratio"  http://dx.doi.org/10.1016/0022-1759(94)90081-7

[lysine]

Hi,

thank you very much for your answer! I will dig into these publications.

lysine