Tim Plant

Serum free light chain measurement aids the diagnosis of myeloma in patients with severe renal failure.

BackgroundMonoclonal free light chains (FLCs) frequently cause rapidly progressive renal failure in patients with multiple myeloma. Immunoassays which provide quantitative measurement of FLCs in serum, have now been adopted into screening algorithms for multiple myeloma and other lymphoproliferative disorders. The assays indicate monoclonal FLC production by the presence of an abnormal κ to λ FLC ratio (reference range 0.26–1.65). Previous work, however, has demonstrated that in patients with renal failure the FLC ratio can be increased above normal with no other evidence of monoclonal proteins suggesting that in this population the range should be extended (reference range 0.37–3.1). This study evaluated the diagnostic sensitivity and specificity of the immunoassays in patients with severe renal failure.MethodsSera from 142 patients with new dialysis-dependent renal failure were assessed by serum protein electrophoresis (SPE), FLC immunoassays and immunofixation electrophoresis. The sensitivity and specificity of the FLC ratios published reference range was compared with the modified renal reference range for identifying patients with multiple myeloma; by receiver operating characteristic curve analysis.ResultsForty one patients had a clinical diagnosis of multiple myeloma; all of these patients had abnormal serum FLC ratios. The modified FLC ratio range increased the specificity of the assays (from 93% to 99%), with no loss of sensitivity. Monoclonal FLCs were identified in the urine from 23 of 24 patients assessed.ConclusionMeasurement of serum FLC concentrations and calculation of the serum κ/λ ratio is a convenient, sensitive and specific method for identifying monoclonal FLC production in patients with multiple myeloma and acute renal failure. Rapid diagnosis in these patients will allow early initiation of disease specific treatment, such as chemotherapy plus or minus therapies for direct removal of FLCs.

Soluble syndecan‐1 level at diagnosis is an independent prognostic factor in multiple myeloma and the extent of fall from diagnosis to plateau predicts for overall survival

Syndecan‐1 (CD138) is a heparin sulphate proteoglycan that is over expressed on the surface of both normal and malignant plasma cells and actively shed from the cell surface (soluble syndecan‐1). Soluble syndecan‐1 has been shown to be an independent prognostic factor in myeloma but its role in prognostic classification requires further investigation. We have retrospectively measured soluble syndecan‐1 in 324 presentation samples and 154 plateau phase samples from the UK Medical Research Council Myeloma VIth trial. Log‐rank analysis showed that the presentation value of soluble syndecan‐1 is a highly significant prognostic factor when assessing survival from entry (χ2 = 14·92, P < 0·0001) and remains an important independent prognostic factor when considered in Cox regression models (P ≤ 0·02) with known independent factors. The magnitude of fall in soluble syndecan‐1 from presentation to plateau also had prognostic value when assessing overall survival from plateau (χ2 = 3·79, P = 0·05). In conclusion, this large study confirms that soluble syndecan‐1 level is a powerful independent prognostic factor both at diagnosis and at plateau phase.

Measurement of antibodies to pneumococcal, meningococcal and haemophilus polysaccharides, and tetanus and diphtheria toxoids using a 19-plexed assay.

The measurement of antibody responses to vaccination is useful in the assessment of immune status in suspected immune deficiency. Previous reliance on enzyme-linked immunoabsorbent assays (ELISA) has been cumbersome, time-consuming and expensive. The availability of flow cytometry systems has led to the development of multiplexed assays enabling simultaneous measurement of antibodies to several antigens. We optimized a flow cytometric bead-based assay to measure IgG and IgM concentrations in serum to 19 antigens contained in groups of bacterial subunit vaccines: pneumococcal vaccines, meningococcal vaccines, Haemophilus influenzae b (Hib), and tetanus and diphtheria toxoid vaccines. 89-SF was employed as the standard serum. The assay was used to determine specific antibody levels in serum from 193 healthy adult donors. IgG and pneumococcal IgM antibody concentrations were measurable across 3 log10 ranges encompassing the threshold protective IgG antibody levels for each antigen. There was little interference between antibody measurements by the 19-plexed assay compared with monoplexed assays, and a lack of cross-reactive IgG antibody, but evidence for cross-reacting IgM antibody for 3/19 pneumococcal antigens. 90th centile values for 15/19 IgG concentrations and 12/12 IgM concentrations of the 193 adult sera were within these ranges and percentages of sera containing protective IgG antibody levels varied from 4% to 95% depending on antigen. This multiplexed assay can simultaneously measure antibody levels to 19 bacterial vaccine antigens. It is suitable for use in standard clinical practice to assess the in vivo immune response to test vaccinations and measure absolute antibody levels to these antigens.

Development of a rapid and quantitative lateral flow assay for the simultaneous measurement of serum κ and λ immunoglobulin free light chains (FLC): inception of a new near-patient FLC screening tool

Abstract Background: Serum free light chains (FLC) are sensitive biomarkers used for the diagnosis and management of plasma cell dyscrasias, such as multiple myeloma (MM), and are central to clinical screening algorithms and therapy response criteria. We have developed a portable, near-patient, lateral-flow test (Seralite®) that quantitates serum FLC in 10 min, and is designed to eliminate sample processing delays and accelerate decision-making in the clinic. Methods: Assay interference, imprecision, lot-to-lot variability, linearity, and the utility of a competitive-inhibition design for the elimination of antigen-excess (‘hook effect’) were assessed. Reference ranges were calculated from 91 healthy donor sera. Preliminary clinical validation was conducted by retrospective analysis of sera from 329 patients. Quantitative and diagnostic results were compared to Freelite®. Results: Seralite® gave a broad competitive-inhibition calibration curve from below 2.5 mg/L to above 200 mg/L, provided good assay linearity (between 1.6 and 208.7 mg/L for κ FLC and between 3.5 and 249.7 mg/L for λ FLC) and sensitivity (1.4 mg/L for κ FLC and 1.7 mg/L for λ FLC), and eliminated anomalous results from antigen-excess. Seralite® gave good diagnostic concordance with Freelite® (Roche Hitachi Cobas C501) identifying an abnormal FLC ratio and FLC difference in 209 patients with newly diagnosed MM and differentiating these patients from normal healthy donors with polyclonal FLC. Conclusions: Seralite® sensitively quantitates FLC and rapidly identifies clinical conditions where FLC are abnormal, including MM.

Brief Report: Immunization of HIV-Infected Adults in the UK With Haemophilus influenzae b/Meningococcal C Glycoconjugate and Pneumococcal Polysaccharide Vaccines.

Abstract:U.K. guidelines for vaccinating HIV-infected adults against bacteria are based on limited data. We compared antibody responses between 211 HIV-infected and 73 HIV-uninfected adults vaccinated with pneumococcal polysaccharide vaccine (PPV) and Haemophilus influenzae b/meningococcal C polysaccharide-tetanus toxoid glycoconjugate vaccine (Hib/MenC-TT). IgG responses to Hib/MenC-TT were not significantly different. PPV induced median IgGs >1.3 &mgr;g/mL for 10/12 serotypes among HIV-uninfected participants and 5/12 in HIV-infected participants. HIV-uninfected adults had higher post-vaccination IgGs than HIV-infected adults for 4/12 serotypes (P < 0.001). Responses did not associate with CD4 count or viral suppression. In a U.K. HIV-infected population, Hib/MenC-TT induced similar responses to HIV-uninfected adults, whereas PPV induced poor responses.

Serum tryptase concentration and progression to end-stage renal disease.

Mast cell activation can lead to nonclassical activation of the Renin–Angiotensin–Aldosterone System. However, the relevance of this to human chronic kidney disease is unknown. We assessed the association between serum tryptase, a product of mast cell activation, and progression to end‐stage renal disease or mortality in patients with advanced chronic kidney disease. We stratified patients by use of angiotensin‐converting enzyme inhibitors/angiotensin receptor II blockers (ACEi/ARB).

What does it mean if a patient is positive for anti-Jo-1 in routine hospital practice? A retrospective nested case-control study

Background: It is widely believed that patients bearing auto-antibodies to histidyl tRNA synthetase (anti-Jo-1) very likely have a connective tissue disease including myositis and interstitial lung disease. The value of positive tests in low disease prevalence settings such as those tested in routine care is unknown. We sought to determine the value of anti-Jo-1 auto-antibodies in routine practice. Methods: Our study was a nested case control study within a retrospective cohort of all patients tested for anti-ENA our hospital, from any hospital department, between January 2013 and December 2014. Data was extracted from electronic records of anti-Jo-1 positive patients and randomly selected ENA negative patients (ratio of 1:2), allowing for a minimum follow up of at least 12 months after first testing. Results: 4009 samples (3581 patients) were tested. Anti-ENA was positive in 616 (17.2%) patients, 40 (1.1%) were anti-Jo-1 positive. Repeat ENA testing was done for 350/3581 (9.8%) patients (428 of 4009 (10.7%) samples) and in 7/40 (17.5%) of anti-Jo-1 positive patients. The median interval between the first and second request was 124 days (inter-quartile range 233 days). The frequencies of interstitial lung disease (ILD), myositis and Raynaud’s were comparable for anti-Jo-1 positive patients (n=40) and 80 randomly selected ENA negative controls. Positive tests led to additional diagnostic testing in the absence of clinical disease. Sensitivity of Jo-1 for ILD was 50% (CI 19-81%), specificity 68% (CI 59-77%), positive predictive value 12.5% (CI 4 to 27%) and negative predictive value 93.8% (CI 86-98%). Of 10 (25%) patients with high anti-Jo1 levels, 3 had ILD, one myositis and two a malignancy (disseminated melanoma and CML). Conclusion: Anti-Jo-1 is uncommon in a heterogenous hospital population and is only weakly predictive for ILD. Repeated test requests were common and potentially unnecessary indicating that controls over repeat requests could yield significant cost savings.