We have examined the role of the murine homologue of Leu-3 T4, L3T4, in recognition of antigen in association with products of the major histocompatibility complex (Ag/MHC) by murine T cell hybridomas. A series of ovalbumin (OVA)/I-Ad-specific T cell hybridomas were ranked in their sensitivity to Ag/I by measuring their ability to respond to low doses of OVA, or their sensitivity to inhibition by anti-I-Ad antibodies. T cell hybridomas with low apparent avidity for OVA/I-Ad, i.e. that did not respond well to low concentrations of OVA and were easily inhibited by anti-I-Ad, were also easily inhibited by anti-L3T4 antibodies. The reverse was true for T cell hybridomas with apparent high avidity for Ag/MHC. We found that the presence of low doses of anti-L3T4 antibodies caused T cell hybridomas to respond less well to low doses of Ag, and to be more easily inhibited by anti-I-Ad antibodies. These results suggested that the role of the L3T4 molecule is to increase the overall avidity of the reaction between T cells and Ag-presenting cells. In support of this idea was the discovery of several L3T4- subclones of one of our L3T4+ T cell hybridomas, D0.11.10. The L3T4- subclones had the same amount of receptor for OVA/I-Ad as their L3T4+ parent, as detected by an anti-receptor monoclonal antibody. The L3T4- subclones, however, responded less well to low doses of OVA, and were more easily inhibited by anti-I-Ad antibodies than their L3T4/ parent. These results showed that the L3T4 molecule was not required for surface expression of, or functional activity of, the T cell receptor for Ag/MHC. The L3T4 molecule did, however, increase the sensitivity with which the T cell reacted with Ag/MHC on Ag-presenting cells.

Background-Transfusion-related acute lung injury (TRALI) is the leading cause of transfusion-related mortality. Blood centers are implementing TRALI risk reduction strategies based on screening apheresis donors for antibodies to human leukocyte antigens (HLA).

A total of 42,160 individuals were typed for HLA-A and HLA-B by both serology and PCR-based typing. The HLA assignments included all of the known serological equivalents. The majority of the individuals (99.9%) were from U.S. minority population groups. The serologic typing was performed between 1993 and 1997 at the time of recruitment for the National Bone Marrow Program (NMDP) registry. The polymerase chain reaction (PCR)-based typing was carried out in two phases. In phase I, DNA typing was performed by PCR using sequence-specific oligonucleotide probes (PCR-SSOP) or PCR using sequence-specific primers (PCR-SSP) without knowledge of the serologic assignments. Discrepancies were identified between the serologic and DNA assignments in 24% of the volunteers (8% of volunteers differed for only HLA-A assignments, 13% for HLA-B, and 3% for both HLA-A and -B) and a potential explanation was assigned each discrepant serology/DNA pair. In phase II, a random sampling scheme was used to select a statistically significant number of individuals for repeat DNA typing from each of these categories. The categories included antigens missed by serology, nonexpressed (null) alleles, PCR amplification failures, misassignment of antigens and nomenclature issues. Only a single individual was found to carry a null allele. DNA-based testing correctly typed nearly 99% of the donors at HLA-A, more than 98% at HLA-B, and more than 97% at both HLA-A and -B validating this methodology for registry typing.

Cells of the gram-negative bacterium Escherichia coli are able to attach to various host cells by means of a mannose-specific adhesin associated with type 1 fimbriae. Here we show that fragmentation of type 1 fimbriae by freezing and thawing results in increased mannose-binding activity as demonstrated by increased hemagglutination, increased stimulation of human lymphocyte proliferation, and increased binding of the mannose-containing enzyme horseradish peroxidase. Increased activity in all three assays was mannose sensitive and was not exhibited by FimH-mutant type 1 fimbriae lacking the adhesin. Scatchard analysis of the data from peroxidase binding assays showed that unfrozen and frozen fimbriae contain binding sites displaying two classes of affinity. Frozen and thawed fimbriae expressed an increase in the number of highaffinity binding sites. These results show that fragmentation of the fimbrial structure exposes cryptic mannosebinding activity associated with type 1 fimbriae, presumably that of internally located adhesin molecules. Our data support earlier observations that adhesin moieties of type 1 fimbriae are located both at the tips and at intervals along the length of the fimbriae. In addition, our data suggest that only the adhesin moieties that are located at the fimbrial tips are functional in binding mannose. Adhesins located along the length of the fimbriae have their mannose-binding activity buried within the fimbrial structure and hence are not functional. We propose an updated model for the structure of type 1 fimbriae that is in agreement with the above observations. Several members of the family Enterobactericeae, including most strains of Escherichia coli, express on their surfaces numerous proteinaceous filaments called fimbriae. The fimbriae expressed by E. coli can be of one or more types based on the specific mode of recognition or interaction mediated by these organelles in promoting adherence of the bacteria to various host cell surfaces (10,12,13,19). In this respect, among the more common fimbriae expressed by E. coli are type 1 fimbriae, which are known to facilitate attachment of the bacteria to mannose-containing receptors exposed on a variety of host cells (4,17,18,22). The mannose-binding activity associated with type 1 fimbriae is commonly demonstrated by specific adhesion assays, including mannose-sensitive agglutination of guinea pig erythrocytes (21). Facilitated adherence of the bacteria to mannose-containing glycoproteins on host cells is believed to be an important prerequisite for successful invasion and colonization in the pathogenesis of several E. coli and other enterobacterial infections (2, 3, 7). Recently, we have found that type 1 fimbriae can also stimulate human lymphocyte proliferation, which is mannose sensitive (20).The structure of type 1 fimbriae was initially described by Brinton as being entirely composed of 17-kDa protein subunits, now It has also been demonstrated that FimH by itself is the mannose-specific adhesin of type 1 fimbriae (11).Although ...

BACKGROUND-TRALI is the leading cause of transfusion-related deaths. Donor HLA antibodies have been implicated in TRALI cases. Blood centers are implementing TRALI risk reduction strategies based on HLA antibody screening of some subpopulations of ever-pregnant apheresis platelet donors. However, if screening assay cutoffs are too sensitive, donation loss may adversely impact blood availability.