Summary report from the first international workshop on soluble HLA antigens. Paris, August 1992

Abstract

The First International Workshop on Soluble HLA antigens focused on the comparison of immunoassay procedures for quantitation of soluble HLA (sHLA) class I antigens and the selection of a sHLA class I antigen international standard. Several sets of serum, plasma, and cell culture supernatant specimens were assayed blindly for levels of sHLA class I antigens by 15 participating laboratories using different immunoassay formats. The sandwich ELISA using (i) for antigen capture: an anti-HLA class I heavy chain monoclonal antibody (mAb) specific for a monomorphic epitope, and (ii) for antigen detection: an anti-beta 2 microglobulin antibody-enzyme conjugate, was the assay format of choice. There was a high inter-laboratory correlation among the majority of laboratories. All serum and plasma specimens from normal donors, and from a single transplant patient, had detectable levels of sHLA class I antigens. Paired serum and plasma specimens had similar levels of sHLA class I antigens, although plasma sHLA antigens seemed more stable than serum sHLA antigens. sHLA-A2 and sHLA-B7 antigens were detected in all specimens from HLA-A2 and HLA-B7 donors, respectively, using allele-specific ELISAs. No difference in reactivity was observed for quantitation of native sHLA class I antigens whether the capture mAb was TP25.99 (alpha 3 domain-specific) or W6/32 (alpha 2 + alpha 3-specific). However, a human-mouse chimeric sHLA class I antigen reacted weakly in assays which used TP25.99 mAb. The wide variation among laboratories in their reporting of micrograms/ml units pointed to the need for an inter-laboratory standardization based on a calibrated sHLA antigen preparation. T.sB7, an sHLA-B7 antigen derived from a cell line transfected within human beta 2 microglobulin and HLA-B7 genes, was accepted as the First sHLA class I Antigen International Standard at the workshop meeting.

Soluble HLA Class I and Class II Concentrations in Factor VIII and PCC Preparations

“…Only small amounts of these molecules could be de tected. Soluble HLA-CI or CII concentrations were always much less than sHLA concentrations found in human plas ma [9,11], In spite of the fact that these preparations are concentrates from pooled plasma, the detected sHLA con centrations were very low compared to the sHLA-CI and sHLA-CII contents found in commercially available IVIG preparations [1,2], It is noteworthy that products No. 5, 6 and 7 did not contain any detectable sHLA-CI or sHLA-CII and only traces of IgG.…”

“…Nunc Maxisorb Immunopiates were coated overnight at sHLA in Factor Vili and PCC Westhoff/Grosse-Wildemostatic preparations sHLA-CII molecules could be de tected (0.02-0.28 mg/1). Compared to the concentrations of sHLA-CI or sHLA-CII in the plasma of healthy individuals (1.01±0.72 mg/1 for sHLA-CI [11] and 1.5±2.4mg/l for sHLA-CII [9], respectively) the concentrations of HLA an tigens in the investigated plasma concentrations was low. The highest sHLA class II concentration was found in prep aration No.…”

“…The measured absorbance (OD: 490 nm) was directly proportional to the amount of sHLA-CI present in the specimens. A reference plasma whose sHLA-CI content had been de termined using the sHLA standard defined by the first international workshop on soluble HLA [11] was included in each assay as a stan dard. The combination of the antibodies used in this modified assay allowed the detection of lower sHLA-CI concentrations than previous ly reported.…”

Immunobiology of Allograft Human Leukocyte Antigens in the New Microenvironment

“…It appears that the level of soluble β2m-free allo-HLA-I is an important factor in the prevention of acute allograft rejection. As mentioned earlier, Hagihara, et al [154], examining sHLA-I changes in pediatric live-donor liver recipients, noted that increased levels of sHLA-I were observed in 10 of 22 (45%) patients without any clinical complications. From the first week to 24 months after grafting, the allograft liver released sHLA-I molecules.…”

“…Hagihara, et al [154], examined sHLA-I changes in 33 pediatric live-donor liver recipients. Three different molecules of sHLA-I (45, 39 and 34-36 kDa) were observed.…”