When “Neat” Just Isnʼt Enough: Gaining Precision and Depth in HLA Antibody Data Interpretation Utilizing Titers and C1q Antibody Binding Assay.

Background: In patients (pts) who are highly sensitized, desensitization therapy is commonly applied to reduce antibody (Abs) load and to reduce potential morbidity and mortality following heart transplant (HTx). It is not known as to what level of antibody binding is needed to initiate desensitization therapy. It is believed that high binding Abs may have the ability to fix complement and lead to cytotoxicity. Methods: Between 2011 and 2012 we assessed 13 highly sensitized pts who were noted to have standardized fluorescence intensity (SFI) >100,000 pretransplant. A complement binding assay (C1q binding assay) was then applied to these Abs to assess for a correlation of binding to cytotoxicity. Correlation statistics were used to derive significance between higher antibody binding and capacity to bind complement. Results: There seems to be no correlation between the different binding strength and C1q positivity. However, when evaluating Luminex 1:8 test, there seems to be correlation between class I binding strength with C1q positivity. There was C1q positive rate of 10.8% in SFI 200,000 range. Conclusions: Undiluted antibodies do not correlate to C1q binding ability. However the 1:8 dilution test seems to provide an better prediction of complement binding. 1:8 dilutions should be performed to accurately assess the ability for antibodies to bind complement.

Download Full-text

Antibodies in Transplantation: The Effects of HLA and Non-HLA Antibody Binding and Mechanisms of Injury

Methods in Molecular Biology - Transplantation Immunology ◽

10.1007/978-1-62703-493-7_2 ◽

2013 ◽

pp. 41-70 ◽

Cited By ~ 47

Author(s):

Nicole M. Valenzuela ◽

Elaine F. Reed

Keyword(s):

Antibody Binding ◽

Hla Antibody

Download Full-text

A Radioimmunometric Antibody-Binding Assay for Evaluation of Xenoantisera to Melanoma-Associated Antigens23

JNCI Journal of the National Cancer Institute ◽

10.1093/jnci/62.3.455 ◽

1979 ◽

Vol 62 (3) ◽

pp. 455-463 ◽

Cited By ~ 26

Author(s):

Richard P. McCabe ◽

Vito Quaranta ◽

Luciano Frugis ◽

Soldano Ferrone ◽

Ralph A. Reisfeld

Keyword(s):

Binding Assay ◽

Antibody Binding

Download Full-text

Improvements to immunoassays by use of covalent binding assay plates

Clinical Chemistry ◽

10.1093/clinchem/40.9.1833 ◽

1994 ◽

Vol 40 (9) ◽

pp. 1833-1837 ◽

Cited By ~ 8

Author(s):

A S Douglas ◽

C A Monteith

Keyword(s):

Alkaline Phosphatase ◽

Edge Effect ◽

Binding Assay ◽

Covalent Binding ◽

Blocking Agent ◽

Antibody Binding ◽

Plate Surface ◽

High Background ◽

Color Development ◽

Capture Antibody

Abstract The uniformity of Xenobind covalent binding microwell plates was compared with that of passive binding plates by binding streptavidin to the plate surface, reacting the streptavidin with biotinylated alkaline phosphatase, and developing color with phenolphthalein monophosphate. Absorbance of the covalent binding plates at 570 nm was 0.95 A (CV 1.9%) compared with 0.35-0.5 A (CV 6.7-16.9%), for passive plates. No edge effect was observed with Xenobind covalent plates. Use of both plates in a model immunoassay gave a CV of 4% for the covalent binding plate and 12-15% for the passive binding plates. Moreover, the background absorbance (color development when no antigen was present) for the covalent binding plates was < 0.05 A in all cases, whereas that for the passive binding plates ranged up to 0.45 A. The high background on passive plates results from displacement of capture antibody and (or) blocking agent from the plate, thereby providing a place for detector antibody binding.

Download Full-text