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Human FAP / Fibroblast activation protein alpha ELISA Kit

SKU:
HUFI02445
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
Q12884
Sensitivity:
0.375ng/ml
Range:
0.625-40ng/ml
ELISA Type:
Sandwich
Synonyms:
FAPalpha, FAP, Seprase, 170 kDa melanoma membrane-bound gelatinase, DKFZp686G13158, DPPIV, EC 3.4.21.-, FAPA, Fibroblast activation protein alpha, fibroblast activation protein, alpha, Integral membrane serine protease, seprase
Reactivity:
Human
€599
Frequently bought together:

Description

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Human FAP/Fibroblast activation protein alpha ELISA Kit

The Human FAP (Fibroblast Activation Protein Alpha) ELISA Kit is a reliable and accurate tool for detecting levels of FAP in human samples such as serum, plasma, and cell culture supernatants. With its high sensitivity and specificity, this kit provides researchers with consistent and reproducible results for a variety of research applications.FAP, also known as fibroblast activation protein alpha, is a key protein involved in tissue remodeling and cancer progression.

Its overexpression has been linked to various types of cancer, making it a valuable biomarker for studying tumor microenvironments and potential therapeutic targets.By using the Human FAP ELISA Kit, researchers can gain valuable insights into the role of FAP in disease progression and explore new avenues for developing targeted therapies. With its ease of use and accuracy, this kit is an essential tool for advancing research in the field of oncology and beyond.

Product Name:Human FAP / Fibroblast activation protein alpha ELISA Kit
Product Code:HUFI02445
Size:96 Assays
Alias:FAPalpha, FAP, Seprase, 170 kDa melanoma membrane-bound gelatinase, DKFZp686G13158, DPPIV, EC 3.4.21.-, FAPA, Fibroblast activation protein alpha, fibroblast activation protein, alpha, Integral membrane serine protease, seprase
Detection method:Sandwich ELISA, Double Antibody
Application:This immunoassay kit allows for the in vitro quantitative determination of Human FAPalpha concentrations in serum plasma and other biological fluids.
Sensitivity:0.375ng/ml
Range:0.625-40ng/ml
Storage:4°C for 6 months
Note:For Research Use Only
Recovery:Matrices listed below were spiked with certain level of Human FAPalpha and the recovery rates were calculated by comparing the measured value to the expected amount of Human FAPalpha in samples.
MatrixRecovery range(%)Average(%)
serum(n=5)92-10399
EDTA plasma(n=5)88-10496
UFH plasma(n=5)85-10295
Linearity:The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human FAPalpha and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
Sample1:21:41:8
serum(n=5)86-97%96-104%85-104%
EDTA plasma(n=5)83-95%86-99%84-101%
UFH plasma(n=5)92-100%82-100%82-99%
CV(%):Intra-Assay: CV<8%
Inter-Assay: CV<10%
ComponentQuantityStorage
ELISA Microplate (Dismountable)8×12 strips4°C for 6 months
Lyophilized Standard24°C/-20°C
Sample/Standard Dilution Buffer20ml4°C
Biotin-labeled Antibody(Concentrated)120ul4°C (Protect from light)
Antibody Dilution Buffer10ml4°C
HRP-Streptavidin Conjugate(SABC)120ul4°C (Protect from light)
SABC Dilution Buffer10ml4°C
TMB Substrate10ml4°C (Protect from light)
Stop Solution10ml4°C
Wash Buffer(25X)30ml4°C
Plate Sealer5-

Other materials and equipment required:

  • Microplate reader with 450 nm wavelength filter
  • Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
  • Incubator
  • Deionized or distilled water
  • Absorbent paper
  • Buffer resevoir
UniprotQ12884
UniProt Protein Function:FAP: In association with DPP4 is involved in the pericellular proteolysis of the extracellular matrix (ECM), the migration and invasion of endothelial cells into the ECM. May have a role in tissue remodeling during development and wound healing, and may contribute to invasiveness in malignant cancers. Belongs to the peptidase S9B family. 2 isoforms of the human protein are produced by alternative splicing.
UniProt Protein Details:Protein type:EC 3.4.14.5; Protease; EC 3.4.21.26; Membrane protein, integral

Chromosomal Location of Human Ortholog: 2q23

Cellular Component: apical part of cell; cell surface; cytoplasm; extracellular space; focal adhesion; integral to membrane; lamellipodium; plasma membrane

Molecular Function:dipeptidyl-peptidase activity; endopeptidase activity; integrin binding; metalloendopeptidase activity; peptidase activity; protease binding; protein binding; protein dimerization activity; protein homodimerization activity; serine-type endopeptidase activity; serine-type peptidase activity

Biological Process: angiogenesis; cell adhesion; endothelial cell migration; proteolysis; proteolysis involved in cellular protein catabolic process; regulation of fibrinolysis

NCBI Summary:The protein encoded by this gene is a homodimeric integral membrane gelatinase belonging to the serine protease family. It is selectively expressed in reactive stromal fibroblasts of epithelial cancers, granulation tissue of healing wounds, and malignant cells of bone and soft tissue sarcomas. This protein is thought to be involved in the control of fibroblast growth or epithelial-mesenchymal interactions during development, tissue repair, and epithelial carcinogenesis. Alternatively spliced transcript variants encoding different isoforms have been found for this gene. [provided by RefSeq, Apr 2014]
UniProt Code:Q12884
NCBI GenInfo Identifier:292495099
NCBI Gene ID:2191
NCBI Accession:Q12884.5
UniProt Secondary Accession:Q12884,O00199, Q53TP5, Q86Z29, Q99998, Q9UID4,
UniProt Related Accession:Q12884
Molecular Weight:
NCBI Full Name:Prolyl endopeptidase FAP
NCBI Synonym Full Names:fibroblast activation protein alpha
NCBI Official Symbol:FAP  
NCBI Official Synonym Symbols:FAPA; SIMP; DPPIV  
NCBI Protein Information:prolyl endopeptidase FAP
UniProt Protein Name:Prolyl endopeptidase FAP
UniProt Synonym Protein Names:170 kDa melanoma membrane-bound gelatinase
Protein Family:Prolyl endopeptidase
UniProt Gene Name:FAP  
UniProt Entry Name:SEPR_HUMAN

*Note: Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.

Before adding to wells, equilibrate the SABC working solution and TMB substrate for at least 30 min at 37°C. When diluting samples and reagents, they must be mixed completely and evenly. It is recommended to plot a standard curve for each test.

StepProtocol
1.Set standard, test sample and control (zero) wells on the pre-coated plate respectively, and then, record their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (zero) wells!
2.Aliquot 0.1ml standard solutions into the standard wells.
3.Add 0.1 ml of Sample / Standard dilution buffer into the control (zero) well.
4.Add 0.1 ml of properly diluted sample ( Human serum, plasma, tissue homogenates and other biological fluids.) into test sample wells.
5.Seal the plate with a cover and incubate at 37 °C for 90 min.
6.Remove the cover and discard the plate content, clap the plate on the absorbent filter papers or other absorbent material. Do NOT let the wells completely dry at any time. Wash plate X2.
7.Add 0.1 ml of Biotin- detection antibody working solution into the above wells (standard, test sample & zero wells). Add the solution at the bottom of each well without touching the side wall.
8.Seal the plate with a cover and incubate at 37°C for 60 min.
9.Remove the cover, and wash plate 3 times with Wash buffer. Let wash buffer rest in wells for 1 min between each wash.
10.Add 0.1 ml of SABC working solution into each well, cover the plate and incubate at 37°C for 30 min.
11.Remove the cover and wash plate 5 times with Wash buffer, and each time let the wash buffer stay in the wells for 1-2 min.
12.Add 90 µl of TMB substrate into each well, cover the plate and incubate at 37°C in dark within 10-20 min. (Note: This incubation time is for reference use only, the optimal time should be determined by end user.) And the shades of blue can be seen in the first 3-4 wells (with most concentrated standard solutions), the other wells show no obvious color.
13.Add 50 µl of Stop solution into each well and mix thoroughly. The color changes into yellow immediately.
14. Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution.

When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.

Sample TypeProtocol
SerumIf using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.

If serum separator tubes are not being used, allow samples to clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.

PlasmaCollect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. Note: Over haemolysed samples are not suitable for use with this kit.
Urine & Cerebrospinal FluidCollect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid.
Cell culture supernatantCollect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately.
Cell lysatesSolubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
Tissue homogenatesThe preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenize in 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or -80°C.
Tissue lysatesRinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
Breast MilkCollect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles.

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