array platforms slide 03-24-2014d1537

2014-08-29

array platforms slide 03-24-2014d1537

slide

some powerpoints this slide was found in
C:\Users\kurtw_000\Documents\kurt\storage\CIM Research Folder\kwhittem\Presentations\2014\PhD Oral Defense\Oral Defense Presentation Kurt Whittemore 4-11-14.pptx

some text from slide 


Summary points


-CIM10K


--17 random, 3 aa linker (GSC)


--aminosilane coated glass surface


--linker on carboxy terminus for CIM10Kv1 and amino terminus for CIM10Kv2


--contact printing with nanoprint for v1 and piezo non-contact printer used by Applied Microarrays for v2


-HT330K


--average length of 12 aa


--similar manufacturing to computer processor chips








Some text from dissertation associated with this info


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Two different random peptide array platforms were used: one platform consisted of an array of 10k peptides (CIM10K) and the other platform consisted of an array of 330k peptides (HT330K).  There were actually two different CIM10K platforms.  The 10k platforms consist of 10k 20 amino acid peptides with sequences of 17 amino acids that were randomly generated, and these peptides covalently bind to a glass slide 40.  Three amino acids of the peptide sequence were not random: glycine, serine, cysteine 41.  These three amino acids formed the linker for attachment to the aminosilane coated glass surface.  This linker is on the carboxyl terminus for CIM10Kv1 and on the amino terminus for CIM10Kv2.  The CIM10Kv1 arrays were produced by spotting peptides synthesized by Alta Biosciences using a NanoPrint LM60 microarray printer (Arrayit, Sunnyvale, CA).  For CIM10Kv2, the peptides were synthesized by Sigma Genosys (St. Louis, MO), and they were printed by Applied Microarrays (Tempe, AZ) using a piezo non-contact printer.


The 330k platform was based on the fabrication of 330k peptide microarrays on a silicon wafer.  This platform also makes use of peptides with a randomly generated sequence.  On this platform, not all of the peptides have exactly the same length, but the average length is 12 amino acids.  The manufacturing process used borrows many techniques from the electronics industry during the production of computer processor chips.  Once the peptides were synthesized onto the silicon wafer, the arrays were deprotectected and soaked overnight in DMF.  The arrays were then stepwise transitioned to an aqueous solution.  The residual DMF was removed by two 5 min washes in distilled water, the arrays were soaked in PBS for 30 min, blocked with an incubation buffer (3% BSA in Phosphate Buffered Saline, 0.05% Tween 20 (PBST)), washed, and then spun dry.  At this point the, the arrays were ready for the application of sera.



Wafer image
http://www.flickr.com/photos/zephinilium/2132084642/sizes/m/

Nanoprint image
image

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