Background: Monitoring of circulating tumor cells (CTCs) in blood of carcinoma patients treated with novel compounds may be a measurement of treatment effectiveness. Before it can be used clinically, a reliably method is needed to enumerate CTCs. We compared two methods for CTC enumeration, OnkoQuick and the CellSearch system. Methods: We drew 22.5 ml of blood into three CellSave tubes from 15 healthy donors and 61 patients with metastatic carcinoma. After pooling, 15 ml was processed with OncoQuick and 7.5 ml with CellSearch.Results: With both methods no CTCs were found in healthy donors. At least one CTC was detected in 14 of 61 patients (23%) with OncoQuick and 33 of 61 patients (54%) with CellSearch (P < 0.0001). The number of CTCs detected was larger for CellSearch (mean 20 CTCs/7.5 ml of blood) than for OncoQuick (3 CTCs/7.5 ml; P < 0.0001).Conclusion: The CellSearch system is a more accurate and sensitive method to enumerate CTCs. Further studies are warranted to evaluate CTC enumeration by the CellSearch system as a monitoring tool for the evaluation of the efficacy of novel anticancer agents. q 2005 International Society for Analytical Cytology

We studied the natural MHC class I display of measles virus (MV) epitopes. Peptide ligands associated with HLA‐A*0201 were purified from a B lymphoblastoid cell line prior to and after infection with MV. Infection‐induced peptides were revealed using microcapillary reversed phase high performance liquid chromatography electrospray ionization / mass spectrometry (μLC‐ESI / MS) by subtraction of the "infected" and "uninfected" ion traces. Three naturally processed viral epitopes derived from different MV proteins were identified through tandem MS sequencing. These peptides were expressed at widely divergent levels of HLA‐peptide complexes, but had similar binding capacities to HLA‐A*0201. The most abundant viral peptide species, identified as residues 84 – 92 (KLWESPQEI) of the MV nonstructural C protein, was expressed at an unprecedented high density (> 105 copies per cell) and was immunogenic in HLA‐A2 / Kb‐transgenic mice. Furthermore, natural mutants of this epitope, occurring in persistent lethal MV strains, were shown to have lost their HLA‐A*0201 binding capacity. Thus, here we report for the first time the direct discovery through μLC‐ESI / MS of a uniquely dominant viral HLA class I ligand, KLWESPQEI, with features eligible for immune selection pressure.

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Herpetic stromal keratitis (HSK) is a T helper type 1 cell-mediated inflammatory disease triggered by herpes simplex virus (HSV) infection of the cornea. In contrast to animal models of HSK, little is known about the role of T cells in human HSK. The phenotypes and repertoires of HSV-specific T cells recovered from the corneas of 12 patients with HSK were determined by flow cytometry. Cornea-derived T cell lines (TCLs) from 10 of the 12 patients contained high numbers of HSV-specific T cells. HSV reactivity was HSV type common and involved relatively more CD8(+) than CD4(+) T cells. The majority of the TCLs showed restricted T cell receptor beta-chain variable region protein (TCRBV) use. T cells expressing 1 or 2 TCRBVs dominated the HSV-1 reactivity in 3 of 5 TCLs analyzed. The data demonstrate that both CD4(+) and CD8(+) T cells may be involved in the HSV-specific T cell response in the corneas of patients with HSK and suggest that restricted TCRBV use by cornea-residing HSV-specific T cells occurs.

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