Development of a novel ELISA to detect total factor D in the presence of high levels of PEGylated anti-factor D Fab

Bioanalysis ◽  
2021 ◽  
Vol 13 (16) ◽  
pp. 1225-1232
Author(s):  
Nancy Yu ◽  
Mehraban Khosraviani ◽  
Yanqiu Liu
Keyword(s):  
Aqueous Humor ◽  
Human Factor ◽  
Drug Tolerance ◽  
Geographic Atrophy ◽  
Alternative Complement Pathway ◽  
Complement Pathway ◽  
Alternative Complement ◽  
High Concentrations ◽  
Rate Limiting ◽  
Commercial Kit

Aim: PEGylated anti-Factor D Fab (PEG-aFD) was developed as a potential therapeutic for geographic atrophy, targeting factor D, the rate-limiting enzyme in the alternative complement pathway. An assay was needed to measure total factor D as a pharmacodynamic biomarker in human aqueous humor in the presence of high PEG-aFD concentrations. Results: Commercial kit met sensitivity requirement, but not drug tolerance requirement. In-house ELISA met both drug tolerance and sensitivity requirements. Addition of 100 ng/ml PEG-aFD to the sample diluent enabled accurate measurement of human factor D in the presence of 2.5 mg/ml of PEG-aFD in the in-house ELISA. Conclusion: Accurate measurement of total factor D in human aqueous humor containing high concentrations of PEG-aFD was achieved by adding PEG-aFD to sample diluent.

Potentiation of factor H by heparin: A rate-limiting mechanism for inhibition of the alternative complement pathway

1983 ◽  
Vol 20 (11) ◽  
pp. 1157-1164 ◽  
Author(s):  
Robert J. Boackle ◽  
Gretchen B. Caughman ◽  
Jana Vesely ◽  
George Medgyesi ◽  
H.Hugh Fudenberg
Keyword(s):  
Factor H ◽  
Alternative Complement Pathway ◽  
Complement Pathway ◽  
Alternative Complement ◽  
Rate Limiting

scholarly journals Simple Method To Distinguish between Primary and Secondary C3 Deficiencies

2003 ◽  
Vol 10 (2) ◽  
pp. 216-220
Author(s):  
Marlene Pereira de Carvalho Florido ◽  
Patrícia Ferreira de Paula ◽  
Lourdes Isaac
Keyword(s):  
Human Serum ◽  
Complement System ◽  
Factor H ◽  
Normal Human Serum ◽  
Alternative Complement Pathway ◽  
Complement Pathway ◽  
Factor B ◽  
Alternative Complement ◽  
Normal Human ◽  
The Complement System

ABSTRACT Due to the increasing numbers of reported clinical cases of complement deficiency in medical centers, clinicians are now more aware of the role of the complement system in the protection against infections caused by microorganisms. Therefore, clinical laboratories are now prepared to perform a number of diagnostic tests of the complement system other than the standard 50% hemolytic component assay. Deficiencies of alternative complement pathway proteins are related to severe and recurrent infections; and the application of easy, reliable, and low-cost methods for their detection and distinction are always welcome, notably in developing countries. When activation of the alternative complement pathway is evaluated in hemolytic agarose plates, some but not all human sera cross-react to form a late linear lysis. Since the formation of this linear lysis is dependent on C3 and factor B, it is possible to use late linear lysis to routinely screen for the presence of deficiencies of alternative human complement pathway proteins such as factor B. Furthermore, since linear lysis is observed between normal human serum and primary C3-deficient serum but not between normal human serum and secondary C3-deficient serum caused by the lack of factor H or factor I, this assay may also be used to discriminate between primary and secondary C3 deficiencies.

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