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NRicher™ C
Enrichment of Complement Cascade Related Proteins
Consumable chemically derived beads, species agnostic as they are not derived from antibodies
Enrich Complement Cascade related proteins from sera/plasma or cell lysates from both animals and humans, >90% Albumin removal
Does not require any specialized instruments, just a standard microfuge
Bead format suitable for automation compatibility, please inquire
On-Bead digestion for LC-MS analysis, or optional elution for any functional, enzymatic, or immunoassay analysis
The complement cascade is a major component of the immune system that provides powerful host surveillance and protection from invading microbes. It also exerts an important influence on the adaptive immune response by acting synergistically with antibodies as well as promoting B- and T-cell stimulation. The intracellularly active complement system—the complosome, has been shown to play a critical function in regulating T cell responses, cell physiology (such as metabolism), and inflammatory disease processes. Complement dysregulation is implicated in chronic diseases ranging from age-related macular degeneration to neurological & cardiovascular disease, as well as cancer. In acute inflammatory conditions, hyperactive complement may predispose individuals to adverse outcomes, as suspected in hospitalized Covid-19 patients.
The complement system consists of over 50 circulating and membrane proteins, comprising about 5% of the total protein mass in plasma. Most complement proteins circulate in blood as inactive precursors (zymogens), that when triggered, become activated through proteolytic cascades. Although textbooks describe three activation pathways (classical, lectin & alternative), largely under-appreciated is Complement’s evolutionarily conserved link to coagulation to eliminate damaged tissues. Complement can activate platelets and contribute to hemostasis in response to injury. Conversely, complement can be activated from proteolytic enzymes derived from coagulation and fibrinolysis that can cleave both C3 and C5 leading to amplification, and final function. As a result of complement activation, many outcomes are produced including: opsonization of pathogens or damaged-self cells to enhance phagocytosis; production of anaphylatoxins C3a & C5a; recruitment of leukocytes to the inflammatory site; and the terminating end of the cascade – assembly of the membrane attack complex (MAC) on the cell surface.
The terminating end of the complement cascade is derived from the C3 Convertase proteolytic product - C5b, which engages the sequential recruitment of C6, C7, C8, and C9, assembling the membrane attack complex (MAC). Also known as the “terminal complement complex”, it results from the coordination of C5b-7 insertion in the membrane, which then captures C8, inducing polymerization of a C9 ring – to as many as 18, C9’s per pore. Terminal MACs perforate the cell membrane of the invading pathogen or target cell, and when a sufficient number of MAC pores form, the cell dies by osmotic lysis. Sub-lytic doses of MAC however, induce dramatically different effects than lytic doses, including adherence, aggregation, chemotaxis, cell division, and extracellular vesicle release.
In such a critical juncture, the complement system must maintain a delicate balance between activation and inhibition to allow activation when necessary to counteract infectious agents or modified self/host tissues, while concurrently protecting healthy self/host tissue. This protection is achieved systemically through the concerted action of regulators and inhibitors ensuring cell and tissue integrity essential for normal and healthy well-being.
NRicher™ C provides enrichment of all Complement-related proteins so as to interrogate functional features derived from proteolysis (split products) or gene variants. Specific tryptic peptides that report functional regions are especially noteworthy after NRicher™ C enrichment; for example - C3a, C4BPB, CFB (Ba fragment), peptides have approximately >10X total spectral signal compared to neat (not enriched) serum. Multiplex targeted proteomics can therefore investigate a more complete picture of Complement activation and regulation using NRicher™ C, than would be possible with immunoassay measurements.
Click Here To View NRicher™ C Product Sheet
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References
"NRicher™
: Family Specific Enrichment For Targeted Proteomics"
Abstract
The
need for new biomarkers to support personalized healthcare, has
fostered numerous proteomic innovations. Still, a number of
challenges remain. One is the preponderance of high abundance
proteins and, concurrently in targeted proteomic workflows,
efficiency and consistency in quantifying target peptides from
different sample cohorts. This is in part due to the changing
landscape of proteins/peptides not associated with the selected
targets. A solution for both these challenges is now available
through a suite of products called NRicher™.
This bead-based technology is derived from experience of over 10
years at the forefront of manufacturing beads (i.e., ionic,
hydrophobic, hydrogen bonding, aromatic, polymeric) with differential
proteome binding properties. NRicher™
consists of consumable chemically derived porous beads, and an
adaptability to bead cocktails, even with seemingly incompatible
surface features; an important distinction of porous, over non-porous
magnetic beads. NRicher™
products do not require any specialized instruments, can be processed
using a standard microfuge, with adaptability to automated liquid
handlers.
Highlights
After
NRicher , target peptides have enriched spectral signal, even as
gradient times are reduced
NRicher
sub-proteome enrichment can minimize acquisition time, collectively
improving overall throughput, cost, and productivity
Investigate
out of the Venn Diagram box. Specific target peptides that report
functional and variant regions promise actionable insights and
potential multiplex biomarkers for disease.
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