Author: Eagle Biosciences

Eagle Biosciences is excited to highlight the Double-Stranded RNA (dsRNA) ELISA Assay Kit from Krishgen Biosystems! This kit allows sensitive and selective detection of dsRNA molecules (larger than 30-40 bp), independent of their nucleotide composition and sequence. Check out the assay background and highlights below!

Background

Double-stranded (ds) RNA formation is a hallmark of viral infections that is essential for the induction of innate immunity. dsRNA is also involve in gene silencing and produced as a side product during in-vitro transcription-based RNA synthesis. The increasing importance of in vitro-transcribed (IVT) mRNA for synthesizing the encoded therapeutic protein in vivo demands the manufacturing of pure mRNA products. The major contaminant in the IVT mRNA is double-stranded RNA (dsRNA). The nuclei of human cells contain dsRNA that plays a role in natural biological processes however, when dsRNA enters the cells, from the extracellular milieu into their endosome or cytoplasm, it is sensed as a viral invader. All cells are capable of responding to dsRNA through sensors and effectors. Therefore, the elimination of dsRNA from the IVT mRNA is crucial to improve translation of the administered mRNA and to limit induction of cytokines.

This assay works on the sandwich-ELISA principle and uses both J2 (IgG2a) antibody to dsRNA and K2 antibody to dsRNA as capture and detection antibodies. dsRNA-recognition is independent of the sequence and nucleotide composition of the antigen.

dsRNA ELISA Kit Highlights

• Pre-coated wells
• Better specificity – using a direct sandwich assay protocol
• Ready-to-Use – all components included, and prepared

If you have any questions about this kit or any of our other offerings, contact us here.

The MedFrontier Intact FGF-23 Assay is developed and manufactured by Minaris Medical! This assay kit is a reliable test that provides a simple and quick assay procedure that yields a broad dynamic range.

What is FGF-23?

FGF-23 (Fibroblast Growth Factor 23) is a protein belonging to the fibroblast growth factor family. FGF-23 is involved in the regulation of phosphorus metabolism. FGF-23 has a molecular weight of approximately 32 kDa. FGF-23 is produced in bone cells. In vivo, FGF-23 is secreted into circulation. A full-length active FGF-23 protein may undergo proteolytic cleavage to generate an inactive c-terminal fragment. The MedFrontier Intact FGF-23 Assay measures ONLY the full-length active form (intact form).

Why Measure FGF-23?

Phosphate plays a critical role in the formation and growth of bones in children and maintaining strength in adults. If there is an imbalance of FGF-23 within the bloodstream, it can cause hyper- and hypophosphatemia. These conditions have been linked to Rickets, osteomalacia, and impaired renal function to name a few.

About the MedFrontier Intact FGF-23 Assay Kit

This is a sandwich Chemiluminescence enzyme immunoassay (CLEIA) kit by using two anti-human FGF-23 mouse monoclonal antibodies. When serum is added to a plate well coated with anti-human FGF-23 mouse monoclonal antibodies, FGF-23 is captured by the immobilized antibodies (1st reaction). After the 1st reaction, the plate is washed. Then, ALP-labeled anti-human FGF-23 mouse monoclonal 2nd antibodies against FGF-23 react to FGF-23 antigens captured by the immobilized antibodies (2nd reaction). After the 2nd reaction, the plate is washed and light detection is performed after adding the luminescence reagent. Each active well of the plate is measured using a luminescence microplate reader and relative light units (RLU) are obtained. The concentration of FGF-23 in serum is calculated with a standard curve generated using the FGF-23 Standard 1 to 6.

Check out the product flyer to learn more!

If you have any questions about this kit or any of our other offerings, contact us here.

We are excited to bring you the newest addition to our line of Complement Assays, two new Factor P assays – one quantitative and one functional. Developed and Manufactured by Svar Life Science, these kits are designed to work optimally together. These assays are ideal for assisting in the development of next-generation complement therapies!

In addition to measuring the properdin (Factor P) concentration and determining its functionality, the Factor P assays can be easily adapted for more in-depth studies, granting a closer look at the amplification loop of the complement system.

About Factor P

Factor P (Complement P, Properdin, FP) is a positive regulator and an initiator of the alternative pathway (AP) for complement activation. It binds surface-bound C3 and C5 convertases and stabilizes them to amplify the activation cascade¹. Factor P binding increases the half-life of the convertase complex approximately 10-fold². It is suggested, however debated, that Factor P also can initiate complement activation by binding for example cell surfaces or certain biological substrates, recruiting C3b or C3(H2O) and Factor B and thus initiate the AP pathway^1,3. Factor P opposes the negative regulation of Factor H that enhances the dissociation of C3b and Bb and mediates Factor I cleavage of C3b to the inactive iC3b3 (Figure 1).

Figure 1: Factor P as stabilizer and initiator of the alternative complement pathway (left) and the opposing negative regulation by Factor H3 (right).

Factor P is not produced in hepatocytes as most complement proteins, but instead by several cell types including monocytes, macrophages, T-cells and granulocytes. It is likely that transient increased concentration of Factor P enhances the AP upon local stimuli4. For example, neutrophils have Factor P-containing granules that are secreted upon stimulation and can enhance the platelet-granulocyte aggregate formation¹. In plasma, Factor P is present in a concentration of approximately 4-25 µg/mL5. Factor P is an elongated 53 kDa glycoprotein that oligomerizes in vivo to dimers, trimers or tetramers (P2, P3 and P4) in a ratio of 26:54:20 (P2:P3:P4) in head to tail structures^5,6. Mutations, deficiencies, protein levels as well as protein deposits of Factor P are connected to diseases and disorders summarized by Chen et al 3. Deficiencies generally increase the susceptibility for meningococcal disease and other infectious diseases7. Altered serum levels have been associated with for example C3 glomerulopathy, Lupus Nephritis, sepsis and chronic heart failure and IgA nephropathy^8.

References

1. Blatt, A. Z.; Pathan, S.; Ferreira, V. P. Properdin: A Tightly Regulated Critical Inflammatory Modulator. Immunol Rev 2016, 274 (1), 172–190. https://doi.org/10.1111/imr.12466.
2. Fearon, D. T.; Austen, K. F. Properdin: Binding to C3b and Stabilization of the C3b-Dependent C3 Convertase. J Exp Med 1975, 142 (4), 856–863.
3. Chen, J. Y.; Cortes, C.; Ferreira, V. P. Properdin: A Multifaceted Molecule Involved in Inflammation and Diseases. Mol. Immunol. 2018, 102, 58–72. https://doi.org/10.1016/j.molimm.2018.05.018.
4. Cortes, C.; Ohtola, J. A.; Saggu, G.; Ferreira, V. P. Local Release of Properdin in the Cellular
   Microenvironment: Role in Pattern Recognition and Amplification of the Alternative Pathway of Complement. Front Immunol 2013, 3. https://doi.org/10.3389/fimmu.2012.00412.
5. Pangburn, M. K. Analysis of the Natural Polymeric Forms of Human Properdin and Their Functions in Complement Activation. The Journal of Immunology 1989, 142 (1), 202–207.
6. Smith, C. A.; Pangburn, M. K.; Vogel, C. W.; Müller-Eberhard, H. J. Molecular Architecture of Human Properdin, a Positive Regulator of the Alternative Pathway of Complement. J. Biol. Chem. 1984, 259 (7), 4582–4588.
7. Fijen, C. A. P.; van den Bogaard, R.; Schipper, M. Properdin Defciency: Molecular Basis and Disease Association. Molecular Immunology 1999, 36, 863–867.
8. Michels, M. A. H. M.; Volokhina, E. B.; van de Kar, N. C. A. J.; van den Heuvel, L. P. W. J. The Role of Properdin in Complement-Mediated Renal Diseases: A New Player in Complement-Inhibiting Therapy? Pediatr Nephrol 2019, 34 (8), 1349–1367. https://doi.org/10.1007/s00467-018-4042-z.

If you have questions about these new items, or any of our other offerings, contact us here.