LabScreen Single Antigen Beads from One Lambda

May 03, 2013

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Stanford University
Structural Biology
Post-Doc

This graph shows the reactivity of 3 KIR-Fc fusion proteins with selected HLA-C allotypes and illustrates well the resolution available with this system.

Company:

One Lambda

Product Name:

LabScreen Single Antigen Beads

Catalog Number:

LS1A04
Image
  • Company: One Lambda
  • Product Name: LabScreen Single Antigen Beads
  • Catalog Number: LS1A04

The One Lambda Labscreen Beads offer a multiplex platform with which to test the specificity of antibodies and recombinant proteins that recognize HLA class I. The panel contains 95 HLA class I allotypes bound to individually identifiable beads. Tests are typically run on a Luminex platform that is able to read the bead identity and the fluorescence of a secondary antibody simultaneously, thus providing a readout of the level of bound antibody/recombinant protein.

Experimental Design and Results Summary

Application

Our lab uses this HLA class I single antigen beads set to assess binding of recombinant killer-cell immunoglobulin like receptor (KIR) to HLA class I allotypes.

Starting Material

Our recombinant proteins are made in an insect cell expression system and solubilized by fusion to the Fc portion of IgG1 antibody. These proteins are used at a concentration of 100ug/ml. For each sample we use 3ul of One Lambda LabScreen single antigen beads.

Protocol Overview

Recombinant Killer-cell Immunoglobulin-like receptor (KIR) proteins are incubated with One Lambda LabScreen single antigen beads for 60 min at 4°C, washed and then stained with an appropriate secondary antibody that recognizes human IgG1 Fc. Samples are analyzed on a Luminex 100 reader that determines the unique identification of a specific bead and the fluorescence of the secondary antibody.

Tips

W6/32: an antibody that recognizes all HLA class I allotypes similarly, can be included in each experiment to normalize for the antigen density on each individual bead.

Results Summary

We see highly specific and highly repeatable results with this beadset. Binding of recombinant KIR proteins is typically between 25 and 90% of that found for W6/32. As an example, KIR2DL1-Fc fusion protein binds highly specifically to only C2+ HLA-C and does not recognize C1+ HLA-C or any HLA-A, HLA-B.

Additional Notes

The peptide content of the HLA class I allotypes on each bead is unknown. However, it is assumed to be highly heterogenous as the HLA class I proteins are derived from EBV transformed B cells.

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Image Gallery

This graph shows the reactivity of 3 KIR-Fc fusion proteins with selected HLA-C allotypes and illustrates well the resolution available with this system.

Summary

The Good

Diverse panel of high quality HLA class I allotypes ideal for use in a cell-free assay.

The Bad

High price, little information on peptide content.

The Bottom Line

A great product that has significantly expanded the ease and scope of KIR/HLA class I interactions.

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