Overview Introduction Workflows Published Data Related Products Related Tests Protocols Q&A Resources
Overview of Complement Factor H
Complement factor H(FH), a plentiful protein found in plasma and serum, serves as a crucial modulator of the complement system's functioning. Its principal role lies in the meticulous control of the complement alternative pathway. In instances of complement activation yielding C3b, interactions occur with factors B and D, culminating in the formation of the C3bBb protease (C3 convertase), a catalyst for further C3 activation and C3b generation.
Fig.1 Complement pathways and the human factor H family.
Introduction of Complement Factor H Functional Test
Why test FH?
FH plays a pivotal role in upholding the balance of an operational complement system, while excessive activation of this system underscores a range of disorders. On the other hand, through its binding to cellular receptors, FH is capable of orchestrating interactions between microbes and host cells. Further investigations will contribute to the assessment of FH's involvement in host-pathogen interactions and the identification of promising new vaccine candidates.
Fig.2 Involvement of the human FH protein family in microbial immune evasion1.
What is the test object?
Serum samples provided by our clients containing either FH or mutant FH can be quantified for concentration using a sandwich ELISA. Subsequently, their functionality will be evaluated through hemolysis assays, encompassing appropriate positive and negative control measures.
Which methods are recommended?
Semi-quantitative enzyme-linked immunosorbent assay (ELISA);
Hemolysis test.
Workflow of FH Functional Test
Sandwich ELISA
Creative Biolabs provides FH detection from plasma/serum, an ELISA can be conducted by employing either a polyclonal or monoclonal antibody for capturing purposes, subsequently utilizing a polyclonal or monoclonal antibody (originating from a distinct species than the capture antibody) for detection. This is followed by the introduction of a secondary antibody enzyme conjugate.
Fig.3 Complement FH ELISA Workflow.
In the functional assessment of FH, a hemolysis test employing sheep erythrocytes is conducted. Serum samples containing either typical FH or variant forms are equally introduced to unsensitized sheep erythrocytes. The extent of sheep erythrocyte lysis is subsequently quantified. Consequently, while normal FH serum does not induce hemolysis in sheep erythrocytes owing to the presence of sialic acid on their surface, serum containing complement factor H variants leads to varying degrees of sheep erythrocyte lysis due to the impaired cell-surface regulatory activity of FH.
Fig.4 Hemolysis test of FH.
Published Data of FH Functional Tests
Insights unveiled in papers elucidating functional assessment of FH:
ELISA analysis
Fig.5 Interaction of FH and factor-H-like protein 1 (FHL-1) with Aspergillus fumigatus enolase (AfEno1).
Fig.6 Binding of FH, FHR1 and FHR5 to glomerular endothelial cells in vitro.
Hemolysis test
Fig.8 Assessment of red blood cell hemolysis in the presence of different factors.
Related Products
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Total Complement Activity Test
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Creative Biolabs offers total complement activity test services for our clients. Total complement activity tests include CH50, AH50, and LP50.
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Creative Biolabs offers high-quality C1q-binding assays for the characterization of therapeutic mAbs.
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Protocols
Resources
Creative Biolabs pioneers a suite of specialized functional assays for diverse complement constituents through cutting-edge technologies and extensive scientific acumen. The complement system is integral to host defense and meticulously regulated in immune responses. FH, a vital modulator of the alternative complement pathway, is among these, and we provide bespoke, top-tier functional tests tailored to our clients' exact requirements, please contact with us for more details.
References
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Jozsi, M. Factor H Family Proteins in Complement Evasion of Microorganisms. Front Immunol. 2017. 8: 571.
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Dasari, P.; et al. Enolase From Aspergillus fumigatus Is a Moonlighting Protein That Binds the Human Plasma Complement Proteins Factor H, FHL-1, C4BP, and Plasminogen. Front Immunol. 2019. 10: 2573.
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Loeven, M. A.; et al. Selective Binding of Heparin/Heparan Sulfate Oligosaccharides to Factor H and Factor H-Related Proteins: Therapeutic Potential for C3 Glomerulopathies. Front Immunol. 2021.12: 676662.
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Dopler, A.; et al. Deregulation of Factor H by Factor H-Related Protein 1 Depends on Sialylation of Host Surfaces. Front Immunol. 2021. 12: 615748.
Questions & Answer
A: An obstacle lies in the intricate nature of FH, a sizable protein engaged in multifaceted interactions, which can render precise functional measurement challenging. Furthermore, disparate assays might yield divergent outcomes, and presently, a universally standardized protocol for conducting FH functional tests is lacking.
A: One avenue entails the formulation of enhanced, universally accepted protocols for complement factor H functional testing, fostering heightened precision and result replicability. Another direction involves delving into the involvement of FH in diverse ailments and circumstances, like age-related macular degeneration, and advancing novel therapeutic approaches targeting the complement system.
A: The complement factor H functional test is a screening tool designed to measure the functional activity of complement factor H in plasma. The test primarily involves the use of specific agents to stimulate the complement system and assess its subsequent response. This test is crucial in immunology and clinical diagnosis research. Researchers can use it to understand the role of complement factor H in immune response, in the pathogenesis of certain diseases like age-related macular degeneration, atypical hemolytic uremic syndrome, and more. It's also used in the development of therapeutic strategies targeting the complement system.
A: Yes, it is possible to conduct multiple tests on a single sample, as long as the volume of the sample is enough. However, this depends on the individual tests requested and the amount of plasma available.
A: Yes. For best results, store plasma samples at -80°C or below and avoid multiple freeze-thaw cycles. For shipping, samples should be sent on dry ice or equivalent frozen condition to preserve the biological integrity of the sample.
A: The test is optimized for plasma, but in some circumstances, it might be possible to use other specimen types. However, additional validation steps might be required, which could impact the turnaround time and cost.
For Research Use Only.
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