Detection of adalimumab and anti-adalimumab antibodies in patients with rheumatoid arthritis: a comprehensive overview of methodology pitfalls and benefits.

About a third of patients with rheumatoid arthritis treated with adalimumab may develop anti-adalimumab antibodies. Anti-adalimumab antibodies are associated with reduced drug levels, loss of drug efficacy, clinical non-response and an increased risk of adverse effects. In case of suspected drug failure and in order to better define clinical efficacy, adalimumab as well as anti-adalimumab antibodies levels should be monitored. Sandwich or indirect enzyme-linked immunoassay is most commonly used for determining adalimumab, while bridging ELISA and antigen-binding test are most useful for determining anti-adalimumab antibodies. Most current assays cannot detect antibodies complexed with the adalimumab; however, methods for dissociation of the complexes using acid/temperature have been developed. The aim of this review is to report on the latest methodology for detecting adalimumab and anti-ADL antibodies, benefits of their detections in clinical practice, as well as expose problematic issues, such as different analytical sensitivity and specificity, standardization and validation. The main problem in measuring adalimumab or anti-ADL antibodies is high drug sensitivity, which can result in false-negative anti-ADL antibodies. Therefore, drug-tolerant assays have been developed. Cell-based assays, such as the reporter gene assay, are recommended for detection of functionally active adalimumab and their neutralizing anti-ADL antibodies.

Metabolic fingerprints of human primary endothelial and fibroblast cells.

INTRODUCTION: Human primary cells originating from different locations within the body could differ greatly in their metabolic phenotypes, influencing both how they act during physiological/pathological processes and how susceptible/resistant they are to a variety of disease risk factors. A novel way to monitor cellular metabolism is through cell energetics assays, so we explored this approach with human primary cell types, as models of sclerotic disorders. OBJECTIVES: In order to better understand pathophysiological processes at the cellular level, our goals were to measure metabolic pathway activities of endothelial cells and fibroblasts, and determine their metabolic phenotype profiles. METHODS: Biolog Phenotype MicroArray™ technology was used for the first time to characterize metabolic phenotypes of diverse primary cells. These colorimetric assays enable detection of utilization of 367 specific biochemical substrates by human endothelial cells from the coronary artery (HCAEC), umbilical vein (HUVEC) and normal, healthy lung fibroblasts (NHLF). RESULTS: Adenosine, inosine, d-mannose and dextrin were strongly utilized by all three cell types, comparable to glucose. Substrates metabolized solely by HCAEC were mannan, pectin, gelatin and prevalently tricarballylic acid. HUVEC did not show any uniquely metabolized substrates whereas NHLF exhibited strong utilization of sugars and carboxylic acids along with amino acids and peptides. CONCLUSION: Taken together, we show for the first time that this simple energetics assay platform enables metabolic characterization of primary cells and that each of the three human cell types examined gives a unique and distinguishable profile.