High sensitivity and specificity of elevated cerebrospinal fluid kappa free light chains in suspected multiple sclerosis.

Cerebrospinal fluid (CSF) analysis is routinely used in the diagnostic work-up of multiple sclerosis (MS), by detecting CSF-specific oligoclonal bands (OCB). More recently, several studies have reported CSF free light chains (FLC) as an alternative. We show that absolute CSF κFLC concentrations were highly sensitive - more than OCB testing - and specific for clinically isolated syndrome, relapsing remitting and primary progressive MS. Measurement of κFLC alone was sufficient. Our results suggest that CSF κFLC levels measured by nephelometry, if validated in a larger series, are a preferred test to OCB analysis in the diagnostic work-up of patients suspected of having MS.

Immunoglobulin heavy/light chain ratios improve paraprotein detection and monitoring, identify residual disease and correlate with survival in multiple myeloma patients.

The novel heavy/light chain (HLC) assay was used for the detection and measurement of monoclonal immunoglobulins, response evaluation and prognostication. This test allows identification and quantification of the different light chain types of each immunoglobulin class (for example, IgGκ and IgGλ) and enables calculation of ratios of monoclonal/polyclonal immunoglobulin (HLC ratio). Sequential sera of 156 patients with IgG or IgA myeloma started on first-line therapy and followed for a median of 46.1 months were analyzed. Results were compared with those obtained with conventional techniques (serum protein electrophoresis (SPEP), immunofixation electrophoresis (IFE), nephelometry (NEPH), and the free light chain test (FLC)). Our data show that the HLC assay allowed quantification of monoclonal proteins not accurately measurable by SPEP or NEPH. When both HLC and FLC testing were applied for response assessment, clonal excess was noted in 14/31 patients with complete response (CR). HLC ratio indicated presence of disease in 8/31 patients who achieved CR and, in sequential studies indicated evolving relapse in three patients before IFE became positive. Highly abnormal HLC ratios at presentation were significantly associated with shorter overall survival (40.5 months vs median not reached, P=0.016). Multivariate analysis revealed HLC ratio (P=0.03) and ß(2)-microglobulin (P<0.01) as independent risk factors for survival.

Memory T cells constitute a subset of the human CD8+CD45RA+ pool with distinct phenotypic and migratory characteristics.

Using HLA class I-viral epitope tetramers to monitor herpes virus-specific CD8(+) T cell responses in humans, we have shown that a significant fraction of responding cells revert from a CD45RO(+) to a CD45RA(+) state after priming. All tetramer-binding CD45RA(+) cells, regardless of epitope specificity, expressed a phenotype LFA-1(high)CCR7(low) that was stable for at least 10 years in infectious mononucleosis patients and indefinitely in asymptomatic carriers. CD8(+)CD45RA(+)LFA-1(high) cells were not present in cord blood but in adults account for up to 50% of CD8(+)CD45RA(+) cells. These CD45RA(+)LFA-1(high) cells have significantly shorter telomeres than CD45RA(+)LFA-1(low) cells, suggesting that the latter represent a naive population, while the former are memory cells. CD45RA(+) memory cells are a stable population of noncycling cells, but on stimulation they are potent producers of IFN-gamma, while naive CD8(+) cells produce only IL-2. The chemokine receptor profile and migratory potential of CD45RA(+) memory cells is very similar to CD45RO(+) cells but different to naive CD8 cells. In accord with this, CD45RA(+) memory cells were significantly underrepresented in lymph nodes, but account for virtually all CD8(+)CD45RA(+) T cells in peripheral tissues of the same individuals.

Quantitative flow cytometry for the analysis of T cell receptor Vbeta chain expression.

Detailed characterisation of the T cell receptor (TCR) repertoire expressed by peripheral blood lymphocytes has been used to study specific T cell responses in disease conditions. The methods have mostly involved molecular biology analysis of transcribed gene products isolated from T cell subsets or individual clones. Extensive characterisation of the TCR Vbeta chain repertoire by flow cytometry is now possible due to the recently increased availability of specific monoclonal antibodies. However, there are major logistical problems inherent in this analysis relating to the number of cells required to obtain accurate results and the vast amounts of data generated. To reduce these factors to a practical level, we have performed a detailed study to define the limits of precision of cell subset analysis by flow cytometry. Maximal achievable precision was obtained by analysing 10(4) lymphocytes; no significant improvement was obtained by analysing greater numbers of cells up to 10(5) cells, even for cell subsets present at frequencies as low as 0.5%. Careful application of these precision profiles will also permit more effective use of clinical research samples for flow cytometry when the availability of cells is limited.