Improvements in microsequencer performance for low picomole sequence analysis Journal Article


Authors: Tempst, P.; Geromanos, S.; Elicone, C.; Erdjument-Bromage, H.
Article Title: Improvements in microsequencer performance for low picomole sequence analysis
Abstract: Five years ago, we reported several simple modifications of the Applied Biosystems 477A-120A sequencing system to improve femtomole PTH-amino acid analysis and to make the process more routine (Anal. Biochem. 183, 290). Since then, several laboratories have obtained extensive sequencing runs with 2-3 picomol initial yield, using either our or their own optimizations. Unfortunately, similar instrument performance has eluded other groups. Optimized ultrasensitive systems can be standard and are easy to operate, even by novice researchers, provided some basic preventive and corrective maintenance routines are strictly adhered to. Here, we review the basics of Edman chemistry, polypeptide sequencing automation, and various limitations inherent to this technique. We describe protocols for (i) peptide preparation and handling, for (ii) improving instrument sensitivity by background reduction and by HPLC optimizations, and for (iii) continued performance assurance. We discuss routine practical operation and what to expect, realistically, from a polypeptide sequencing facility. © 1994 Academic Press. All rights reserved.
Journal Title: Methods
Volume: 6
Issue: 3
ISSN: 1046-2023
Publisher: Academic Press Inc., Elsevier Science  
Date Published: 1994-09-01
Start Page: 248
End Page: 261
Language: English
DOI: 10.1006/meth.1994.1027
PROVIDER: scopus
DOI/URL:
Notes: Export Date: 14 January 2019 -- Article -- Source: Scopus
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  1. Paul J Tempst
    307 Tempst