Synthesis of Random Peptides for Antibody Purification 5-17-12

2015-01-13

azim58 - Synthesis of Random Peptides for Antibody Purification 5-17-12

I'll pick the top 10 most tumor significant peptides for synthesis from
the Application of 4T1 and Naive Sera onto Random Peptide Array 4-26-12
experiment.

These peptides will be synthesized onto the Tentagel S NH2 (90 micron
beads). Donnie mentioned that he thinks Bart usually does a small 0.05 mM
scale. I should input the peptides that I want to use into the GEMS
database.

Sequence of the 6 random peptides I want synthesized

Name
 tumor
 tumor
 naïve
 naïve
 p value

CSGYNPIWHVHEFSMNYPTK
 591
 590
 251
 250
 4.33E-06

CSGQESKYTYPDDFNAKYTE
 203
 202
 151
 152
 0.000192

CSGVNGEYSMYFSASHWGTH
 238
 240
 149
 147
 0.000241

CSGNRVWWYTKQDGWHFLRD
 1006
 997
 527
 521
 0.000348

CSGNKNATDYSTPHIEIMET
 161
 160
 123
 124
 0.000365

CSGKHAWNIPKRPPFDQNDG
 6459
 6604
 1409
 1278
 0.000379

Note that all of these peptides are higher in the tumor than the naive.

original data here:
"L:\storage\CIM Research Folder\DR\2012\5-14-12\ran pep
analysis\4T1_and_naive_raw_good_spots.xls"
from experiment here
http://www.azim58.wikispaces.net/Application+of+4T1+and+Naive+Sera+onto+Ran
dom+Peptide+Array+4-26-12

Mara checked the purity of these peptides from the mass spec data here
"S:\Research\Cancer_Eradication\Discovering tumor specific
antigens\Antibody purification\purification of random peptides with sera
6-11-12\Six Peptide Mass Spec Purity.doc"

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6-4-12 received 1st peptide from Donnie "K-1 2409 MW" ->
CSGYNPIWHVHEFSMNYPTK stored at 4 C in box with dessicator bag
6-5-12 received 2nd peptide from Donnie "K-2 2345 MW" ->
CSGQESKYTYPDDFNAKYTE stored at 4 C in box with dessicator bag

===========================================================================
7-2-12 Requested that peptides be synthesized at 4X the density on the
Tentagel beads.

Here is my e-mail to Donnie:
Hey Donnie, I talked to Kathy and she does want me to have those peptides
synthesized at a higher density on Tentagel beads. I've reinput the 6
sequences back into the GEMS software (also below). I would want the
peptides so that two lysines are closest to the surface of the beads, and
then the CSG. . . part of the peptide goes out from the lysines away from
the bead. The two lysines at the beginning should be blocked with Fmoc
and Fmoc rather than Fmoc and Boc. The purpose of this is to cause 4
peptide sequences to be created from these two lysines to increase the
density of the peptides. Let me know if any of that is unclear or if I am
confused about anything. Thanks!

CSGYNPIWHVHEFSMNYPTK
CSGQESKYTYPDDFNAKYTE
CSGVNGEYSMYFSASHWGTH
CSGNRVWWYTKQDGWHFLRD
CSGNKNATDYSTPHIEIMET
CSGKHAWNIPKRPPFDQNDG

Donnie plans to start synthesizing these peptides on Thursday (7-12-12)

sketch from Donnie with two lysines
double lysine sketch from Donnie.pdf

7-13-12 There was a change to these synthesis plans. After talking with
Zhao, we realized that the amino acids would be linked differently even
if the order away from the surface was kept the same. Therefore, Donnie
plans to synthesize these peptides (not on a bead), Alex will purify the
peptides, and then these peptides will be conjugated to the Tentagel
surface.
see Orientation of Peptides Synthesized onto Tentagel Beads

8-2-12 These peptides have now been synthesized: 8-2-12 Peptides

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