Abstract:
A project has been initiated to identify proteins in the blood of cancer patients. Plasma,
one of the major components of blood, has been chosen for this study as it is easily
separated. In addition it consists of a large number of proteins compared to any other
tissue, cell or organ. Therefore, the plasma proteome is an ideal source of diagnostic
markers and therapeutic targets for many human diseases.
In this study, One Dimensional Sodium Dodecyl Sulphate Polyacrylamide Gel
Electrophoresis (1D – SDS - PAGE) coupled with Matrix Assisted Laser Desorption
Ionization Time Of Flight (MALDI-TOF) Mass Spectrometry (MS) has been used to
identify plasma proteins. In order to investigate the feasibility of this technique, plasma
from healthy individuals has been used. Plasma samples were electrophoresized under
denaturing conditions and silver staining was carried out to visualize proteins. A total of
40-50 gel protein bands were detected in healthy individuals and of these, selected protein
bands were excised, trypsinized and then subjected to MALDI-TOF-MS analysis. The
identification of proteins was performed by peptide mapping using sequence databases.
The mass of peptides derived from an in-gel proteolytic digestion were measured and
subsequently searched against the protein databases available on internet.
By using the above technique, the major plasma proteins, Transferrin, Macroglobulin
alpha 2, Albumin, Proapolipoprotein and a few other plasma proteins were identified with
sequence coverage range of 16 – 54 %. It has been possible to obtain sequences of all the
identified proteins as well as their few modifications. This technique, already being
employed at the Department of Physics, University of Colombo, could be applied to
identify plasma proteins in cancer patients in Sri Lanka by comparing their protein profiles
with those of healthy persons. The results of this work clearly indicate that the particular
technique has the potential as a tool of clinical investigations.
