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

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+Random+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"

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

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