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- Analysis report

Date: 2021-09-22

Submitted by: Isabelle Sirois

Analysis type: Class II

Description of experiment:

MHC CLASS II MOUSE A20

Samples:

MHC_CLASS_II_FROM_MOUSE_A20_CELLS_-_M5_ANTIBODY: Alleles: H-2-IAd, H-2-IEd

Species: MOUSE A20

# of cells: 100 M

Lysis buffer: 0.5 % CHAPS

Type of beads: 80 mg Sepharose CNBr-activated beads

Antibody: M5 - H2 IAd/IEd - 2 MG

Incubation time: overnight

MHC-ligand complex elution buffer: 1% TFA

Peptide elution buffer: 28% ACN/0.1% TFA

Type of MS/MS: ORBITRAP ECLIPSE TRIBID MASS SPECTROMETER

Peptide identification software: PEAKS PRO V10.6

Peptide FDR: 1%

Sample Overview

- LF Score: fraction of peptides between 9 and 22 mers. - BF Score: fraction of peptides between 9 and 22 mers which are predicted to be strong or weak binders.

Sample Total peptides Peptides between 9-22 mers LF Score BF Score
MHC_CLASS_II_FROM_MOUSE_A20_CELLS_-_M5_ANTIBODY 2295 2219 0.97 0.87

Peptide Length Distribution (maximum of 30 mers)

Annotation Results

NetMHCIIpan eluted ligand predictions made for all peptides between 9 & 22 mers, inclusive. - Percent rank cutoffs for strong and weak binders: 2.0 and 10.0. - Percentages are calculated across rows (i.e. percentage of total peptides for a respective sample).

Sample Total peptides Allele Strong binders Weak binders Non-binders

MHC_CLASS_II_FROM_MOUSE_A20_CELLS_-_M5_ANTIBODY

 2219 H-2-IAd 956 (43.1%) 338 (15.2%) 925 (41.7%)
H-2-IEd 556 (25.1%) 514 (23.2%) 1149 (51.8%)
Binding Affinities

Binding Heatmaps

NetMHCIIpan eluted ligand predictions made for all peptides between 9 & 22 mers, inclusive. Approximate color legend (detailed mapping shown next to heatmaps):

Predicted strong binders (%rank <= 2.0)

Predicted weak binders (2.0 < %rank <= 10.0)

Predicted non-binders

- - # of peptides in sample

Sequence Motifs

Clustering performed with all peptides between 9 & 22 mers, inclusive.

- Percentages represent the percentage of peptides in a given group predicted to strongly bind the indicated allele.

Polar

Neutral

Basic

Acidic

Hydrophobic

MHC_CLASS_II_FROM_MOUSE_A20_CELLS_-_M5_ANTIBODY (peptides clustered: 2219, outliers: 101)

Peptides in group: 792

H-2-IAd: 6%,

H-2-IEd: 55%

Peptides in group: 1326

H-2-IAd: 69%,

H-2-IEd: 9%

MHC_CLASS_II_FROM_MOUSE_A20_CELLS_-_M5_ANTIBODY sequence motif(s)
H-2-IAd

Peptides: 1277

H-2-IEd

Peptides: 1046

Non-binders group 2

Peptides: 140

Non-binders group 1

Peptides: 116