Mouse FGA (Fibrinogen Alpha chain) ELISA Kit (MOES01017)

SKU:
MOES01017
€549

Description

ELISA Kit Technical ManualMSDS

Mouse FGA (Fibrinogen Alpha chain) ELISA Kit

This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Mouse FGA . Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Mouse FGA and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Mouse FGA, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Mouse FGA. The concentration of Mouse FGA in samples can be calculated by comparing the OD of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityMouse
Detection MethodColormetric
Detection Range31.25-2000 ng/mL
Sensitivity18.75 ng/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Mouse FGA in samples. No significant cross-reactivity or interference between Mouse FGA and analogues was observed.

Typical Data

As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e.g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.

Concentration
(ng/mL)
O.D Average Corrected
20002.378
2.43
2.4042.328
10001.569
1.627
1.5981.522
5000.904
0.9
0.9020.826
2500.428
0.452
0.440.364
1250.266
0.25
0.2580.182
62.50.192
0.162
0.1770.101
31.250.122
0.134
0.1280.052
00.072
0.08
0.076--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Mouse FGA were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Mouse FGA were tested on 3 different plates, 20 replicates in each plate.

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (ng/mL)95.03264.80985.30102.59286.05895.28
Standard deviation6.3011.9732.816.2416.6834.47
C V (%)6.634.523.336.085.833.85

Recovery

The recovery of Mouse FGA spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.

Sample Type Range (%) Average Recovery (%)
Serum (n=5)84-9791
EDTA plasma (n=5)94-107100
Cell culture media (n=5)86-9891

Linearity

Samples were spiked with high concentrations of Mouse FGA and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.

Serum (n=5) EDTA plasma (n=5) Cell culture media (n=5)
1:2Range (%)87-10187-10390-102
Average (%)929497
1:4Range (%)88-10382-9586-99
Average (%)968791
1:8Range (%)87-9985-9981-96
Average (%)949288
1:16Range (%)87-10079-9387-101
Average (%)928592

Kit Components & Storage

An unopened kit can be stored at 4'C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.

Item Specifications Storage
Micro ELISA Plate(Dismountable)8 wells X12 strips-20'C, 6 months
Reference Standard2 vials
Concentrated Biotinylated Detection Ab (100X)1 vial, 120 uL
Concentrated HRP Conjugate (100X)1 vial, 120 uL-20'C(shading light), 6 months
Reference Standard & Sample Diluent1 vial, 20 mL4'C, 6 months
Biotinylated Detection Ab Diluent1 vial, 14 mL
HRP Conjugate Diluent1 vial, 14 mL
Concentrated Wash Buffer (25X)1 vial, 30 mL
Substrate Reagent1 vial, 10 mL4'C(shading light)
Stop Solution1 vial, 10 mL4'C
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy

Mouse FGA (Fibrinogen Alpha chain) ELISA Kit (MOES01017) Assay procedure

    1. Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
    2. Aliquot 100µl of standard solutions into the standard wells.
    3. Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
    4. Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
    5. Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
    6. Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix.Incubate for 1 hour at 37°C.
    7. Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
    8. Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
    9. Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
    10. Add 90µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37°C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
    11. Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
    12. Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.

Mouse FGA (Fibrinogen Alpha chain) ELISA Kit (MOES01017) Protein Information

UniProt Protein Function:FGA: Fibrinogen has a double Function: yielding monomers that polymerize into fibrin and acting as a cofactor in platelet aggregation. Defects in FGA are a cause of congenital afibrinogenemia (CAFBN). This is a rare autosomal recessive disorder characterized by bleeding that varies from mild to severe and by complete absence or extremely low levels of plasma and platelet fibrinogen. The majority of cases of afibrinogenemia are due to truncating mutations. Variations in position Arg-35 (the site of cleavage of fibrinopeptide a by thrombin) leads to alpha- dysfibrinogenemias. Defects in FGA are a cause of amyloidosis type 8 (AMYL8); also known as systemic non-neuropathic amyloidosis or Ostertag-type amyloidosis. AMYL8 is a hereditary generalized amyloidosis due to deposition of apolipoprotein A1, fibrinogen and lysozyme amyloids. Viscera are particularly affected. There is no involvement of the nervous system. Clinical features include renal amyloidosis resulting in nephrotic syndrome, arterial hypertension, hepatosplenomegaly, cholestasis, petechial skin rash. 2 isoforms of the human protein are produced by alternative splicing.
UniProt Protein Details:

Protein type:Secreted; Secreted, signal peptide

Chromosomal Location of Human Ortholog: 3 E3|3 36.96 cM

Cellular Component: blood microparticle; cell cortex; cell surface; cytoplasm; external side of plasma membrane; extracellular exosome; extracellular region; extracellular space; fibrinogen complex; rough endoplasmic reticulum

Molecular Function:cell adhesion molecule binding; metal ion binding; protein binding, bridging; receptor binding; structural molecule activity

Biological Process: adaptive immune response; blood coagulation; blood coagulation, common pathway; blood coagulation, fibrin clot formation; cell-matrix adhesion; cellular protein complex assembly; fibrinolysis; hemostasis; immune system process; induction of bacterial agglutination; innate immune response; negative regulation of endothelial cell apoptotic process; negative regulation of extrinsic apoptotic signaling pathway via death domain receptors; plasminogen activation; platelet activation; positive regulation of ERK1 and ERK2 cascade; positive regulation of exocytosis; positive regulation of heterotypic cell-cell adhesion; positive regulation of peptide hormone secretion; positive regulation of protein secretion; positive regulation of smooth muscle cell migration; positive regulation of vasoconstriction; protein complex assembly; protein polymerization; response to calcium ion; signal transduction

NCBI Summary:This gene encodes a subunit of the coagulation factor fibrinogen, which is a component of the blood clot. The encoded protein is proteolytically processed by thrombin during the conversion of fibrinogen to fibrin. Mice lacking the encoded protein display bleeding in the peritoneal cavity, skin and soft tissues around joints immediately after birth, and are predisposed to spontaneous fatal abdominal hemorrhage as they grow. Pregnant mice lacking the encoded protein succumb to uterine bleeding during gestation. Alternative splicing results in multiple transcript variants encoding different isoforms that may undergo similar proteolytic processing. [provided by RefSeq, Nov 2015]
UniProt Code:E9PV24
NCBI GenInfo Identifier:167555029
NCBI Gene ID:14161
NCBI Accession:NP_001104518.1
UniProt Secondary Accession:E9PV24,Q99K47,
UniProt Related Accession:E9PV24
Molecular Weight:61,326 Da
NCBI Full Name:fibrinogen alpha chain isoform 1 preproprotein
NCBI Synonym Full Names:fibrinogen alpha chain
NCBI Official Symbol:Fga  
NCBI Official Synonym Symbols:Fib  
NCBI Protein Information:fibrinogen alpha chain
UniProt Protein Name:Fibrinogen alpha chain
Protein Family:Fibrinogen
UniProt Gene Name:Fga  
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Additional Information

Product type:
ELISA
Reactivity:
Mouse
ELISA Type:
Sandwich
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