19%, 7.99%, and 6.96%, respectively. Assay batch-to-batch variability was assessed by analysing 50 serum samples with varying FLC levels (κ range 3.42–329.88 mg/L; λ range 1.09–130.51 mg/L) click here and the results are displayed in Fig. 7. All samples were analysed once, on separate assay days, using three consecutive batches of anti-FLC mAbs, calibrators and other appropriate assay reagents. Passing and Bablok regression analysis gave slopes between 0.93–1.01 for κ FLC and 0.86–1.05 for λ FLC. Spearman correlation coefficients for κ FLC were

≥ 0.99 and for λ FLC were ≥ 0.96. Representative assay linearity results are displayed in Fig. 8. Serum samples containing high levels of either κ (581.36, 416.37, and 256.97 mg/L) or λ (485.04, 379.41and 370.56 mg/L) FLC paraproteins were serially diluted in assay buffer. Results indicated that assay linearity was maintained on the monoclonal κ FLC samples between 7.61 mg/L and 568.01 mg/L, 1.94 mg/L and 410.36 mg/L, and, 6.32 mg/L and 260.78 mg/L, respectively. For the λ monoclonal FLC samples, linearity was maintained between 1.38 mg/L and 476.1 mg/L, 1.78 mg/L and 361.72 mg/L, and, 4.45 mg/L and 381.62 mg/L, respectively. For κ FLC, below 10 mg/L no more than 1.45 mg/L non-linearity was found, and above 10 mg/L no more than 16.37% non-linearity was observed. For λ FLC, below 10 mg/L no

more than 2.03 mg/L non-linearity was found, selleck chemical and above 10 mg/L no more than 19.0% non-linearity was found. The assay limit of detection Lepirudin for each mAb was assessed by measuring each against a κ or λ BJ protein, firstly mixed with normal serum, and then

serially diluted in assay buffer. Limit of detection for BUCIS 01 was 0.63 mg/L, BUCIS 04 was 0.86 mg/L, BUCIS 03 was 0.72 mg/L, and BUCIS 09 was 0.52 mg/L. Assay interference tests showed minimal assay cross-reactivity to alternate κ or λ FLC or intact immunoglobulins, bilirubin, haemoglobin, cholesterol or triglyceride (Fig. 9, in supplementary data). Results demonstrated that no more than a median 2.7 mg/L change was observed for the anti-κ FLC mAbs, and no more than a median 3.7 mg/L change for the anti-λ FLC mAbs. This study describes the development of four mouse anti-human κ:λ FLC mAbs and their initial validation in a multi-plex Luminex® immunoassay. Each of the anti-FLC mAbs exhibited: excellent sensitivity (< 1 mg/L); low batch variation; sustained assay linearity; specificity and minimal cross-reactivity to bound LC, or alternate FLC isotype. Each of the mAbs provided good quantitative concordance with the Freelite™ assay in the measurement of polyclonal FLC in plasma from 249 healthy donors, and FLC levels in serum from 1000 consecutive samples. Specificity and sensitivity were further illustrated in the measurement of FLC in 13,090 urine samples tested for BJ proteins.