Biology; Chemistry and chemical biology; Chemical and biological engineering; Biomedical engineering
In Copyright : this Item is protected by copyright and/or related rights. You are free to use this Item in any way that is permitted by the copyright and related rights legislation that applies to your use. For other uses you need to obtain permission from the rights-holder(s). https://rightsstatements.org/page/InC/1.0/;
Two-Dimension Affinity Resolution Electrophoresis Demonstrates that Three Distinct Heparin Populations Interact with Antithrombin III R. E. Edens, J. R. Fromm, S. J. Fromm, R.J. Linhardt, J. M. Weiler, Biochemistry, 34, 2400-2407, 1995.
Tween 20 is frequently used in blotting analysis of proteins and nucleic acids to decrease background staining due to nonspecific binding of antibodies to various membranes. Inclusion of 0.1% (w/v) Tween 20 in the washing buffer (Buffer 1) significantly decreased the chemiluminescence signals of both LMW-EGF and HMW-EGFs (data not shown). This result indicates that the binding of proteins to PVDF membranes is susceptible to the detergent even after fixation with formaldehyde. In conclusion, we suggest the following conditions for the analysis ofLMW-EGF: (a) treat PVDF membranes with a 500 J.tg/ml gelatin solution, (b) carry out electroblotting for 10 min (the duration depends on the blotting conditions used), (c) fix the membranes with formaldehyde after blotting, and (d) do not include Tween 20 in the washing buffer. The optimized conditions made it possible to detect 0.1-0.3 ng of rat LMWEGF (Fig. 3). AJthough the conditions are not optimum for the detection of HMW-EGFs, as described above, it is possible to analyze LMW- and HMW-EGF species simultaneously on a single membrane with this method.;
Biochemistry, 34, 2400-2407; Note : if this item contains full text it may be a preprint, author manuscript, or a Gold OA copy that permits redistribution with a license such as CC BY. The final version is available through the publisher’s platform.
The Linhardt Research Labs.; The Shirley Ann Jackson, Ph.D. Center for Biotechnology and Interdisciplinary Studies (CBIS);
The Linhardt Research Labs Online Collection; Rensselaer Polytechnic Institute, Troy, NY; https://harc.rpi.edu/;