A direct immunoassay assessment of streptavidin- and N-hydroxysuccinimide-modified biochips in validation of serological TNFalpha responses in hemophagocytic lymphohistiocytosis.
The authors report 2 biochip platforms on gold manufactured by either nanoscale biotinylated self-assembled architectures to streptavidin surface or proteins containing free NH(2) groups to N-hydroxysuccinimide (NHS)-activated surfaces and investigated the potential application of tumor necrosis factor-alpha (TNFalpha) serodiagnosis of hemophagocytic lymphohistiocytosis (HLH). Interactions of TNFalpha antigen and TNFalpha antibody on the biochips were optimized using an indirect immunofluorescence method. Variation coefficients were 1.87% to 4.56% on the streptavidin biochip and 5.03% to 8.64% on the NHS biochip. The correlation coefficients (r) in TNFalpha and TNFalpha antibody assays in HLH patients between the 2 biochip formats were 0.9623 and 0.9386 and the concordance frequencies were 92.2% and 96.1%, respectively. To detect plasma TNFalpha-receptor complexes (TNFR1 and R2) in HLH, a biochip assay strategy was developed. Plasma levels of TNFalpha, TNFalpha antibody, and TNFalpha-receptor complexes (TNFR1 and R2) were detected in plasmas from 42 HLH cases using streptavidin biochips. Frequencies of the biomarkers in the plasmas were 40.5% (17/42) for TNFalpha, 30.9% (13/42) for TNFalpha antibody, 28.6% (12/42) for TNFalpha-receptor 1 complex, and 26.1% (11/42) for TNFalpha-receptor 2 complex, respectively. The streptavidin biochip format was more sensitive than the NHS surface and was demonstrated to be a valuable tool to identify individual biomarker molecules and molecular complexes in sera and cell lysates and to track therapeutic progress of patients.
Weidong Du
Sektion Experimentelle Anaesthesiologie, Universitaetsklinikum Ulm, Ulm, Germany. weidong.du@uni-ulm.de
Xueling Ma
Schneider
E Marion
EM
D000937:Antigen-Antibody Reactions / Q000276:immunology; D015415:Biomarkers / Q000097:blood; D019921:Biotinylation; D020099:Coated Materials, Biocompatible / Q000138:chemical synthesis* / Q000737:chemistry; D004492:Edetic Acid / Q000494:pharmacology; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D005456:Fluorescent Dyes / Q000378:metabolism; D006046:Gold; D006801:Humans; D007118:Immunoassay; D051359:Lymphohistiocytosis, Hemophagocytic / Q000175:diagnosis*; D008954:Models, Biological; D040081:Protein Array Analysis / Q000295:instrumentation / Q000379:methods*; D011994:Recombinant Proteins / Q000378:metabolism; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012698:Serologic Tests; D019809:Streptavidin / Q000737:chemistry*; D013388:Succinimides / Q000737:chemistry*; D013499:Surface Properties; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism
D015415:Biomarkers; D020099:Coated Materials, Biocompatible; D005456:Fluorescent Dyes; D011994:Recombinant Proteins; D013388:Succinimides; D014409:Tumor Necrosis Factor-alpha; D006046:Gold; D019809:Streptavidin; D004492:Edetic Acid; D016650:Fluorescein-5-isothiocyanate; C001426:N-hydroxysuccinimide