Sickle SCAN™ (BioMedomics) fulfills analytical conditions for neonatal screening of sickle cell disease.

Sickle SCAN™ is a rapid, qualitative, point-of-care lateral flow immunoassay for the identification of AS, AC, SS/Sß0thal, SC and CC/Cß0thal phenotype. We evaluated this test under the conditions encountered in the French newborn screening (NBS) program for sickle cell disease: a total of 104 dried blood spots (DBSs) were tested with an HPLC reference method and then with the Sickle SCAN™ device. Sickle SCAN™ identified the hemoglobin (Hb) phenotype correctly on 96% of cases. In the four non-concordant cases, the antibody anti-HbS cross-reacted with HbE (n=2), HbD (n=1) or HbX (n=1). There were no false negative. In order to test Sickle SCAN™'s sensitivity to low levels of HbA and HbS in the presence of high HbF levels, we selected another 21 DBS cards with low percentages of HbA (0.6-4.2%) and HbS (2.0-6.9%). HbA and HbS were always detected when present at levels of more than 1% and 2%, respectively. Sickle SCAN™ appears to be an accurate point-of-care method for the identification of newborns with SCD trait. The device meets the criteria for sickle cell disease NBS programs in endemic countries with poor access to laboratory equipment.

A Minimum Epitope Overlap between Infections Strongly Narrows the Emerging T Cell Repertoire.

Many infections are caused by pathogens that are similar, but not identical, to previously encountered viruses, bacteria, or vaccines. In such re-infections, pathogens introduce known antigens, which are recognized by memory T cells and new antigens that activate naive T cells. How preexisting memory T cells impact the repertoire of T cells responding to new antigens is still largely unknown. We demonstrate that even a minimum epitope overlap between infections strongly increases the activation threshold and narrows the diversity of T cells recruited in response to new antigens. Thus, minimal cross-reactivity between infections can significantly impact the outcome of a subsequent immune response. Interestingly, we found that non-transferrable memory T cells are most effective in raising the activation threshold. Our findings have implications for designing vaccines and suggest that vaccines meant to target low-affinity T cells are less effective when they contain a strong CD8 T cell epitope that has previously been encountered.

Autoimmunity-associated LYP-W620 does not impair thymic negative selection of autoreactive T cells.

A C1858T (R620W) variation in the PTPN22 gene encoding the tyrosine phosphatase LYP is a major risk factor for human autoimmunity. LYP is a known negative regulator of signaling through the T cell receptor (TCR), and murine Ptpn22 plays a role in thymic selection. However, the mechanism of action of the R620W variant in autoimmunity remains unclear. One model holds that LYP-W620 is a gain-of-function phosphatase that causes alterations in thymic negative selection and/or thymic output of regulatory T cells (Treg) through inhibition of thymic TCR signaling. To test this model, we generated mice in which the human LYP-W620 variant or its phosphatase-inactive mutant are expressed in developing thymocytes under control of the proximal Lck promoter. We found that LYP-W620 expression results in diminished thymocyte TCR signaling, thus modeling a "gain-of-function" of LYP at the signaling level. However, LYP-W620 transgenic mice display no alterations of thymic negative selection and no anomalies in thymic output of CD4(+)Foxp3(+) Treg were detected in these mice. Lck promoter-directed expression of the human transgene also causes no alteration in thymic repertoire or increase in disease severity in a model of rheumatoid arthritis, which depends on skewed thymic selection of CD4(+) T cells. Our data suggest that a gain-of-function of LYP is unlikely to increase risk of autoimmunity through alterations of thymic selection and that LYP likely acts in the periphery perhaps selectively in regulatory T cells or in another cell type to increase risk of autoimmunity.

Autoreactive T cells bypass negative selection and respond to self-antigen stimulation during infection.

Central and peripheral tolerance prevent autoimmunity by deleting the most aggressive CD8(+) T cells but they spare cells that react weakly to tissue-restricted antigen (TRA). To reveal the functional characteristics of these spared cells, we generated a transgenic mouse expressing the TCR of a TRA-specific T cell that had escaped negative selection. Interestingly, the isolated TCR matches the affinity/avidity threshold for negatively selecting T cells, and when developing transgenic cells are exposed to their TRA in the thymus, only a fraction of them are eliminated but significant numbers enter the periphery. In contrast to high avidity cells, low avidity T cells persist in the antigen-positive periphery with no signs of anergy, unresponsiveness, or prior activation. Upon activation during an infection they cause autoimmunity and form memory cells. Unexpectedly, peptide ligands that are weaker in stimulating the transgenic T cells than the thymic threshold ligand also induce profound activation in the periphery. Thus, the peripheral T cell activation threshold during an infection is below that of negative selection for TRA. These results demonstrate the existence of a level of self-reactivity to TRA to which the thymus confers no protection and illustrate that organ damage can occur without genetic predisposition to autoimmunity.