Human Glycocalicin ELISA kit

Instructions
Code CSB-E12101h
Size 96T,5×96T,10×96T
Trial Size 24T ELISA kits trial application
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Product Details

Description

CUSABIO’s Human Glycocalicin ELISA kit is designed for the quantitative detection of human glycocalicin in serum, plasma, or tissue homogenates. This kit has been validated with high sensitivity, excellent specificity, good linearity, high recovery, precision low than 10%, and high lot-to-lot consistency. The detection mechanism of this kit is based on the sandwich ELISA technique and enzyme-substrate chromogenic reaction. The solution color develops in proportion to the amount of glycocalicin in the sample, and the intensity of the color can be measured at 450 nm via a microplate reader.

Platelet glycocalicin is the major and extramembranous domain of Glycoprotein Iba (GPlba) that can be rapidly cleaved by proteases such as calpain, plasmin, and trypsin. It is not only a marker but also a parameter that reflects platelet turnover, therefore being an aid in the classification of thrombocytopenic disorders. Besides, platelet glycocalicin could act as a circulating inhibitor of thrombin. The levels of platelet glycocalicin were elevated in patients with myocardial infarction after receiving recombinant tissue plasminogen activator (rtPA) treatment. Platelet glycocalicin levels in platelet concentrates may be helpful to qualitative control the storage conditions over time for blood banks.

Target Name glycoprotein Ib (platelet), alpha polypeptide
Alternative Names GP1BA ELISA Kit; Platelet glycoprotein Ib alpha chain ELISA Kit; GP-Ib alpha ELISA Kit; GPIb-alpha ELISA Kit; GPIbA ELISA Kit; Glycoprotein Ibalpha ELISA Kit; Antigen CD42b-alpha ELISA Kit; CD antigen CD42b) [Cleaved into: Glycocalicin] ELISA Kit
Abbreviation GP1BA
Uniprot No. P07359
Species Homo sapiens (Human)
Sample Types serum, plasma, tissue homogenates
Detection Range 0.156 μg/mL-10 μg/mL
Sensitivity 0.039 μg/mL
Assay Time 1-5h
Sample Volume 50-100ul
Detection Wavelength 450 nm
Research Area Blood Coagulation
Assay Principle quantitative
Measurement Sandwich
Precision
Intra-assay Precision (Precision within an assay): CV%<8%
Three samples of known concentration were tested twenty times on one plate to assess.
Inter-assay Precision (Precision between assays): CV%<10%
Three samples of known concentration were tested in twenty assays to assess.
Linearity
To assess the linearity of the assay, samples were spiked with high concentrations of human glycocalicin in various matrices and diluted with the Sample Diluent to produce samples with values within the dynamic range of the assay.
 SampleSerum(n=4)
1:1Average %90
Range %87-94
1:2Average %95
Range %91-99
1:4Average %84
Range %80-88
1:8Average %86
Range %82-90
Recovery
The recovery of human glycocalicin spiked to levels throughout the range of the assay in various matrices was evaluated. Samples were diluted prior to assay as directed in the Sample Preparation section.
Sample TypeAverage % RecoveryRange
Serum (n=5) 8984-93
EDTA plasma (n=4)9287-96
Typical Data
These standard curves are provided for demonstration only. A standard curve should be generated for each set of samples assayed.
μg/ml. OD1OD2AverageCorrected
102.246 2.268 2.257 2.130
51.395 1.374 1.385 1.258
2.50.766 0.798 0.782 0.655
1.250.488 0.462 0.475 0.348
0.6250.326 0.354 0.340 0.213
0.3120.220 0.233 0.227 0.100
0.1560.168 0.164 0.166 0.039
00.126 0.128 0.127  
Materials provided
  • A micro ELISA plate ---The 96-well plate has been pre-coated with an anti-human glycocalicin. This dismountable microplate can be divided into 12 x 8 strip plates.
  • Two vials lyophilized standard ---Dilute a bottle of the standard at dilution series, read the OD values, and then draw a standard curve.
  • One vial Biotin-labeled glycocalicin antibody (100 x concentrate) (120 μl/bottle) ---Act as the detection antibody.
  • One vial HRP-avidin (100 x concentrate) (120 μl/bottle) ---Bind to the detection antibody and react with the TMB substrate to make the solution chromogenic.
  • One vial Biotin-antibody Diluent (15 ml/bottle) ---Dilute the Biotin-antibody.
  • One vial HRP-avidin Diluent (15 ml/bottle) ---Dilute the HRP-avidin solution.
  • One vial Sample Diluent (50 ml/bottle)---Dilute the sample to an appropriate concentration.
  • One vial Wash Buffer (25 x concentrate) (20 ml/bottle) ---Wash away unbound or free substances.
  • One vial TMB Substrate (10 ml/bottle) ---Act as the chromogenic agent. TMB interacts with HRP, eliciting the solution turns blue.
  • One vial Stop Solution (10 ml/bottle) ---Stop the color reaction. The solution color immediately turns from blue to yellow.
  • Four Adhesive Strips (For 96 wells) --- Cover the microplate when incubation.
  • An instruction manual
Materials not provided
  • A microplate reader capable of measuring absorbance at 450 nm, with the correction wavelength set at 540 nm or 570 nm.
  • An incubator can provide stable incubation conditions up to 37°C±5°C.
  • Centrifuge
  • Vortex
  • Squirt bottle, manifold dispenser, or automated microplate washer
  • Absorbent paper for blotting the microtiter plate
  • 50-300ul multi-channel micropipette
  • Pipette tips
  • Single-channel micropipette with different ranges
  • 100ml and 500ml graduated cylinders
  • Deionized or distilled water
  • Timer
  • Test tubes for dilution
Troubleshooting
and FAQs
ELISA kit FAQs
Storage Store at 2-8°C. Please refer to protocol.
Lead Time 3-5 working days

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Target Background

Function
(From Uniprot)
GP-Ib, a surface membrane protein of platelets, participates in the formation of platelet plugs by binding to the A1 domain of vWF, which is already bound to the subendothelium.
Gene References into Functions
  1. An autosomal dominant mode of inheritance, a family history of mild bleeding episodes, aggregation pattern in affected individuals together with evidence of mutation occurring in part of the GP1BA gene encoding the leucine-rich repeat region suggest a novel variant causing monoallelic Bernard-Soulier syndrome. PMID: 30332551
  2. Combined deficiency of factors V and VIII by chance coinheritance of parahaemophilia and haemophilia A, but not by mutations of either LMAN1 or MCFD2 PMID: 29119711
  3. A review of mutations associated with Bernard-Soulier Syndrome and platelet type von Willebrand disease (review). PMID: 28961024
  4. ERK5 associates with CKII to play essential roles in GPIb-IX-mediated platelet activation via the PTEN/PI3K/Akt pathway. PMID: 28603902
  5. There was no evidence to suggest that polymorphisms of GP VI T13254C and GP Ibalpha VNTR were associated with CAD. PMID: 28607925
  6. analysis of an artificial botrocetin that can inhibit the VWF-GPIb interaction PMID: 28071872
  7. Loss of the platelet surface receptors GPIbalpha and GPVI in heart failure, CF-VAD and ECMO patients may contribute to ablated platelet adhesion/activation, and limit thrombus formation under high/pathologic shear conditions PMID: 27601054
  8. Very low birth weight preterm neonates have increased numbers of platelets interacting with von Willebrand Factor, and increased GPIbalpha expression on the platelet surface PMID: 27416003
  9. Data suggest that an aspartate at position 1261 is the most critical residue of VWF N-terminal linker for inhibiting binding of VWF A1 domain to GP1BA on platelets in a model simulating blood flow velocity; network of salt bridges between Asp1261 and rest of VWF A1 domain lock N-terminal linker in place such that binding to GP1BA is reduced. PMID: 28924049
  10. Data show that von Willebrand factor (VWF) is first converted from a compact to linear form by flow, and is subsequently activated to bind platelet glycoprotein Ib alpha polypeptide (GPIbalpha) in a tension-dependent manner. PMID: 28831047
  11. The >30 nm macroglycopeptide separating the two domains of GPIbalpha transmits force on the VWF-GPIbalpha bond (whose lifetime is prolonged by leucine-rich repeat domain unfolding) to the juxtamembrane mechanosensitive domain to enhance its unfolding, resulting in unfolding cooperativity at an optimal force. PMID: 27434669
  12. Meta-analysis found that glycoprotein Ia C807T T allele or the TT genotype, the Ser-allele of HPA-3 and B allele of glycoprotein Ibalpha variable number tandem repeat polymorphisms were associated with increased risk for ischemic stroke. PMID: 28004990
  13. Our results suggest that the -5CC genotype in Kozak sequence of GPIb-alpha may be associated with a higher risk of developing arterial ischemia of lower limbs in type 2 diabetes mellitus patients. PMID: 27888791
  14. Specific inhibition of GPIbalpha shedding in the stored platelets improves post-transfusion platelet recovery and hemostatic function, providing clear evidence for GPIbalpha shedding as a cause of platelet clearance. PMID: 27417583
  15. miR-10a and miR10b regulate the expression of human platelet GP1BA and GP1bb for normal megakaryopoiesis. PMID: 27834869
  16. Data indicate that binding of hemoblobin (Hb) to glycoprotein1balpha (GP1balpha) induced platelet activation plays a crucial role in thrombus formation on immobilized von Willebrand factor (VWF) or type I collagen under shear stresses. PMID: 27105433
  17. Lateral dimerization of GPIbalpha induced by antibody binding is not sufficient to initiate GPIb-IX signaling and induce platelet clearance. PMID: 26662889
  18. GPIb alpha plays a critical role in the co-localization of thrombin and factor XI and the resultant efficient activation of FXI PMID: 12968031
  19. Hemoglobin interaction with GP1balpha induces platelet activation and apoptosis: a novel mechanism associated with intravascular hemolysis. PMID: 26341739
  20. Both GPIbalpha and PAR4 are required for thrombin-induced reactive oxygen species formation PMID: 26569550
  21. Our results reveal the molecular mechanism of collagen-regulated, A1-mediated platelet adhesion enhancement. PMID: 26213126
  22. Report no relationship, between polymorphisms of platelet membrane glycoprotein Ibalpha and risk of coronary heart disease in Chinese Han population. PMID: 26191334
  23. There is a prevalence of Thr145Met and T(-5)C GP Iba polymorphiams in patients with atherotrombotic stroke due to macroangiopathy. PMID: 26539867
  24. Molecular analysis demonstrated a novel homozygous c.800C>G substitution in GP1BA exon 2 leading to a serine 267 Ter stop codon in all 3 siblings PMID: 26044173
  25. Whereas VWF-D'D3 is the major regulator of soluble VWF binding to platelet GpIbalpha, both the D'D3-domain and N-terminal peptide regulate platelet translocation and thrombus formation. PMID: 25341886
  26. Data indicate that GPIbalpha clustering induced by anti-GPIbalpha N-terminus antibody causes integrin alphaIIbbeta3-dependent platelet aggregation, phagocytosis, and rapid platelet clearance in the liver. PMID: 25231551
  27. Data show that force can switch the kinetics of bond formation between A1 domain of von Willebrand factor (VWF) and glycoprotein Ibalpha (GPIbalpha). PMID: 25810255
  28. novel mutation that causes von Willebrand disease type 2B is identified in a German family PMID: 24337418
  29. The platelet adhesion receptor, the glycoprotein Ib-IX-V complex, not only mediates platelet adhesion but also transmits signals leading to platelet activation, aggregation and secretion PMID: 17414217
  30. data indicated that GPIIb-IIIa and GPIb levels are mainly affected by platelet size (MPV) but not by their genetic variations; in some acute coronary syndrome patients, production of large platelets with high GPIIb-IIIa and GPIb contents might be stimulated by elevated thrombopoietin PMID: 23941967
  31. Data indicate a G > T in platelet glycoprotein Ib alpha polypeptide (GP1BA) gene, resulting in a Trp to Leu amino acid change at residue 246 (p.W246L), and this mutation was absent in his unaffected mother and also in the 100 controls. PMID: 24474090
  32. clone 5G6 showed similar inhibitory potency as a widely used shedding inhibitor GM6001 in both constitutive and induced GPIbalpha shedding in human platelets PMID: 24119228
  33. VWF interaction with glycoprotein Ib is modified by polyphosphate PMID: 23006049
  34. Our findings indicate that ADAM17 may be a risk factor for ischemic stroke in Chinese and the expression of GPIbalpha can serve as a measure for stroke severity. PMID: 23771674
  35. Platelet GP Ib-IX can be considered a multifunctional participant in hemostasis, thrombosis, and the inflammatory cascade. PMID: 24504734
  36. Increased mean platelet volume values correlated with increased platelet aggregation activity and enhanced GP IIb-IIIa and GP Ib expression. PMID: 24749250
  37. These findings suggest that structural changes, including central GPIbalpha LRR-A1 contact, contribute to VWF affinity regulation. PMID: 24391089
  38. genetic association study in population in western India: Data suggest novel mutations in platelet glycoprotein Ib (GP1BA, GP1BB) and GP9 are associated with Bernard-Soulier syndrome in subjects studies; of 12 mutations identified, ten were novel. PMID: 23995613
  39. Studies indicate that the platelet-type von Willebrand disease (PT-VWD) is caused by gain-of-function mutations in the platelet GP1BA gene, which codes for the platelet von Willebrand factor (VWF) receptor, GPIbalpha. PMID: 23934752
  40. Studies indicate that platelets from Bernard-Soulier syndrome (BSS) are defective in glycoprotein (GP)Ib-IX-V, a platelet-specific adhesion-signaling complex, composed of GPIbalpha disulfide linked to GPIbbeta, and noncovalently associated with GPIX and GPV. PMID: 23929303
  41. signaling process through the GPIbalpha cytoplasmic tail required for full platelet activation is defective in BSS variant case II and a length polymorphism of GPIbalpha is associated with a modified level of RIPA heterozygous BSS case I. PMID: 23414566
  42. analysis of the localized dynamics-driven affinity regulation mechanism for vWF-GPIbalpha interaction PMID: 23902764
  43. These data indicate an important role for the platelet adhesion receptor GPIb-IX in endotoxin-induced thrombosis and thrombocytopenia. PMID: 24051142
  44. A considerable fraction of the Jordanian population is resistant to the antiplatelet effect of aspirin, which might be related to GPIba C-5T polymorphism. PMID: 23688555
  45. A GP1BA exon 2 fragment spanned the HPA-2 polymorphism and adjacent sequences. PMID: 23750933
  46. genotype may predict the development of septic emboli in patients with infective endocarditis PMID: 23611001
  47. identify a novel Asp235Tyrmutation in the GP1BA gene of two Iranian patients showing the PT-VWD phenotype who were originally misdiagnosed as type 2B VWD PMID: 23014764
  48. GPIX increased the expression of GPIba by promoting the formation of a disulfide bond between GPIba and GPIbb in transfected CHO-K1 cells. PMID: 23143686
  49. Data indicate that exposure of von Willebrand factor sites for glycoprotein Ibalpha binding and ADAMTS13 cleavage are coupled. PMID: 22922961
  50. The T1255A, Clus1, and DC variants caused increased ristocetin-mediated GPIbalpha binding to VWF. PMID: 22517896

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Involvement in disease Non-arteritic anterior ischemic optic neuropathy (NAION); Bernard-Soulier syndrome (BSS); Bernard-Soulier syndrome A2, autosomal dominant (BSSA2); Pseudo-von Willebrand disease (VWDP)
Subcellular Location Membrane, Single-pass type I membrane protein
Database Links

HGNC: 4439

OMIM: 153670

KEGG: hsa:2811

STRING: 9606.ENSP00000329380

UniGene: Hs.1472

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