BDB -BiopanningDataSet Details

Biopanning Data

BiopanningDataSet ID: 3267

Peptides:

Sequences	Affinity Values
CIGNSNTLC(38.4%) CTVRTSADC(23.2%) CSNNGNALC(15.4%) CISNGNQPC(15.4%) CRVNTAALC(7.6%)	[6.29] [15.4] [47.3] [64.2] [77.7]

Sequences Features:

Motif:

Unique Sequence Number:

Experimental Method:

Phage display (common panning)

Sequencing Platform:

Sequencing Method:

Sanger sequencing

Rounds of Panning:

Reference:

PMID: 29029605

Target Name:

Immunoglobulins (Igs) of murine 5T33MM cells

Template Name:

Not determined.

Structure of Target-Template Complex:

Not determined.

Structure of Target-Peptide Complex:

Not determined.

Library Name:

Ph.D.-C7C phage display library (CX7C)

Affinity Measurement Method:

ELISA

Affinity Measurement Description:

The specific binding of peptides to the 5T33MM sIgG was analyzed by ELISA. Streptavidin coated 96 well plates were washed extensively and supplemented with biotin-conjugated peptides by 1 h-incubation at 37 °C; after washing and blocking with blocking solution (1X PBS, 0.05% Tween-20, 5% milk), aliquots of 5T33MM sIgG were added overnight at 4 °C. Wells were extensively washed and coated with the anti-mouse IgG (Fc-specific) alkaline phosphatase-conjugated [Sigma Aldrich] for 1 h at 37 °C, incubated with the alkaline phosphatase substrate, and analyzed by an ELISA reader at 405 nm [Labsystems multiscan MS]. KD values (nM) of identified synthetic peptides are shown.

Brief Description:

The screening of phage displayed library was performed using the bait 5T33MM Igs. Briefly, the streptavidin-conjugated beads were coated with 5T33MM Igs and incubated with 1.0e11 phages overnight at 4 °C. 5T33MM Igs-interacting phages were eluted with 0.2 M glycine-HCl (pH 2.2, 1 mg/mL BSA) followed by the addition of neutralizing solution (1 M Tris-HCl pH 9.1). Ultimately, 3 cycles of panning were performed. Plaques of lysis from isolated phages were transferred to nitrocellulose filters, and membranes were blocked with PBS 1X, 0.1% NP-40; 5% milk; 0.02% NaN3 and then incubated for 2 h at RT with 100 μg of purified 5T33MM Igs. After washing, membranes were hybridized with an alkaline phosphatase-conjugated anti-mouse IgG (dilution 1:5000) and then washed 6 times. Immunoreactive phage clones were detected by BCIP/NBT premixed substrate.

BiopanningDataSet Comments:

Based on the highest affinity binding to 5T33MM Igs, the insert amino acid sequence of phage clone 5 (CIGNSNTLC) was used for large-scale synthesis of the p5 peptide. By FACS, the FITC-conjugated P5 detected the MM-released exosomes in the serum of 5T33MM-engrafted mice, levels of which are correlated with tumor progression at an earlier time point compared to serum paraprotein.

Curator:

Lixu Jiang

Curation Date:

2018-01-19

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