Fibrinogen Assay Kit | FGG assay kit

Rat Fibrinogen ELISA assay

Cat. Num. AAA480412

• Synonyms: Fibrinogen; Rat Fibrinogen ELISA assay; Rat Fibrinogen Antigen Assay; FGG assay kit

• Sequence Length: 444

• Procedural Notes: 1. This assay should be run at room temperature.; 2. Use aseptic technique when opening and dispensing reagents.; 3. This kit is designed to work properly as provided and instructed. Additions, deletions or substitutions to the procedure or reagents are not recommended, as they may be detrimental to the assay.; 4. To minimize error due to handling, wipe the exterior bottom of the microplate wells with a lint-free paper towel.
• Reagent Preparation: 1. 10x Wash Buffer: Dilute the 50 mL of concentrate to 1x with 450 mL of DI water prior to use.; 2. 5x Dilution Buffer: Dilute the 50 mL of concentrate to 1x with 200 mL of DI water prior to use.; 3. Primary Antibody: Reconstitute with 1x Dilution Buffer as directed on the vial and vortex gently to mix. Prepare immediately prior to use.; 4. Secondary Conjugate: Dilute with 1x Dilution Buffer as directed on the vial and vortex gently to mix. Prepare immediately prior to use.
• Standard Preparation: Reconstitute the standard by adding 5.0 mL of dilution buffer directly to the vial and agitate to fully dissolve. This will provide a concentration of 800 ng/mL solution. Prepare the serial dilution as outlined in the table below.; Standard Fibrinogen Concentration (ng/mL) Amount of 1x Dilution Buffer (muL) Transfer Volume (muL) Transfer Source Total Volume (muL); S9 800 --- 1000 Stock Vial 500; S8 400 500 500 S9 500; S7 200 500 500 S8 500; S6 100 500 500 S7 500; S5 50 500 500 S6 500; S4 25 500 500 S5 500; S3 12.5 500 500 S4 500; S2 6.25 500 500 S3 500; S1 3.125 500 500 S2 500; S0 0 500 ---- --- 500
• Sample Collection, Storage and Preparation: Samples giving rat fibrinogen levels above 800 ng/ml should be diluted in 1X Dilution Buffer before use. Normal plasma samples need to be diluted between 1:10,000 and 1:50,000 in 1X Dilution Buffer for the values to be within the linear range of the standard curve.
• Preparation and Storage: 1. Store the kit and all of its components at 4 degree C before use.; 2. If not using the entire plate at once, prepare only the appropriate amount of Primary Antibody and Standard. The remaining stock solutions should be frozen and stored at At -70 degree C. Do not freeze/thaw more than once. All other components should remain refrigerated.; 3. Store unused portions of the microplate in a pouch with a desiccant at 4 degree C.

Typical Testing Data/Standard Curve (for reference only)

Related Product Information for FGG assay kit

Introduction: Fibrinogen is a soluble glycoprotein that circulates in the blood and is converted to insoluble fibrin by thrombin in the final step of the coagulation cascade 1. Hepatic expression of fibrinogen increases two to four hundred fold during the acute phase response to infection or inflammation
2. Elevated fibrinogen levels are correlated with cardiovascular disease3 and atherosclerosis4.

Principle of assay: This is an ELISA (Enzyme-Linked Immunosorbent Assay) for the quantitative analysis of total fibrinogen levels in rat plasma and serum. This test kit operates on the basis of sandwich ELISA where the fibrinogen binds to a capture antibody on the plate and is quantified with the use of a biotin labeled primary antibody and an avidin-HRP conjugate.
Rat fibrinogen will bind to the affinity purified capture antibody coated on the microtiter plate. After appropriate washing steps, biotin labeled polyclonal anti-rat fibrinogen primary antibody binds to the captured protein. Excess antibody is washed away and bound polyclonal antibody is reacted with avidin conjugated to horseradish peroxidase. TMB substrate is used for color development at 450nm. A standard calibration curve is prepared along with the samples to be measured using dilutions of rat fibrinogen. Color development is proportional to the concentration of fibrinogen in the samples.

References

1. Kamath, S. and Lip, G.Y.H.; (2003) QJM 96:711-729
2. Kusher, I.; (1982) Ann New York Acad Sci 389:39-48
3. Kannel, W.B., et al.; (1987) J Am Med Assoc 258:1183-1186
4. Handa, K., et al.; (1989) Atherosclerosis 77:209-213
5. Larrson, A., et al.; (1997) Vet Immunol Immunopathol 59:163-169
6. Dorner, H., et al., (1995) Gerontology 41:252-259

NCBI and Uniprot Product Information

• NCBI GI #: 350
• NCBI GeneID: 280792
• UniProt Accession #: P12799
• Molecular Weight: 50,244 Da
• NCBI Official Full Name: fibrinogen
• NCBI Official Symbol: FGG
• NCBI Protein Information: fibrinogen gamma-B chain; fibrinogen, gamma polypeptide; gamma'
• UniProt Protein Name: Fibrinogen gamma-B chain
• Protein Family: Fibrinogen
• UniProt Gene Name: FGG
• UniProt Synonym Gene Names: Gamma'
• UniProt Entry Name: FIBG_BOVIN

Uniprot Description

Together with fibrinogen alpha (FGA) and fibrinogen beta (FGB), polymerizes to form an insoluble fibrin matrix. Has a major function in hemostasis as one of the primary components of blood clots. In addition, functions during the early stages of wound repair to stabilize the lesion and guide cell migration during re-epithelialization. Was originally thought to be essential for platelet aggregation, based on in vitro studies using anticoagulated blood. However, subsequent studies have shown that it is not absolutely required for thrombus formation in vivo. Enhances expression of SELP in activated platelets via an ITGB3-dependent pathway. Maternal fibrinogen is essential for successful pregnancy. Fibrin deposition is also associated with infection, where it protects against IFNG-mediated hemorrhage. May also facilitate the antibacterial immune response via both innate and T-cell mediated pathways.