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Human TIMP1 / TIMP ( EPA / CLGI ) ELISA Kit
 
 
 
 
 
 
 
Order or Inquire for Human TIMP1 / TIMP ( EPA / CLGI ) ELISA Kit
 
 
 
 
 
 
High affinity Human TIMP1 / TIMP ( EPA / CLGI ) ELISA Kit
 
 
 
 
 
 
Detection limit - 3.125 pg/ml
 
 
 
 
 
 
Affordable price and 30%-80% cost saving for Bulk order
 
 
 
 
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Materials
 
Protocol
 
Trouble Shooting
 
Image & Flash
 
 
 
 
1. Materials provided
 
Capture Antibody - 1.0 mg/mL of mouse anti-TIMP1 monoclonal antibody. Dilute to a working concentration of 2.0 μg/mL in CBS before coating.
 
Detection Antibody - 0.5 mg/mL of biotinylated rabbit anti-TIMP1 polyclonal antibody. Dilute to a working concentration of 1.0 μg/mL in detection antibody dilution buffer before use.
 
Standard - Each vial contains 4.8 ng of recombinant TIMP1. Reconstitute standard powder with detection antibody dilution buffer. After reconstitution, store at -20℃ to -70℃ in a manual defrost freezer. A seven-point standard curve using 2-fold serial dilutions in sample dilution buffer, and a high standard of 200 pg/mL is recommended.
 
Streptavidin-HRP - 50 μL of streptavidin conjugated to horseradish-peroxidase. 1:2000 Dilution in detection antibody dilution buffer before use.
 
 
 
2. Sensitivity
 
The minimum detectable dose of human TIMP1 ( EPA / CLGI ) was determined to be approximately 3.125 pg/ml. This is defined as at least three times standard deviations above the mean optical density of 10 replicates of the zero standard.
 
 
 
3. Principle of the product
 
The human TIMP1 ( EPA / CLGI ) ELISA kit is for the quantitative determination of human TIMP1.
 
This ELISA kit contains the basic components required for the development of sandwich ELISAs. Each kit contains sufficient materials to run ELISAs on five 96-well plates.
 
The Sino Biological ELISA kit is a solid phase sandwich ELISA (Enzyme-Linked Immunosorbent Assay). It utilizes a monoclonal antibody specific for TIMP1 ( EPA / CLGI ) coated on a 96-well plate. Standards and samples are added to the wells, and any TIMP1 ( EPA / CLGI ) present binds to the immobilized antibody. The wells are washed and a biotinylated rabbit anti- TIMP1 polyclonal antibody is then added, producing an antibody-antigen-antibody “sandwich”. To produces color in proportion to the amount of TIMP1 ( EPA / CLGI ) present in the sample strepavidin-HRP and TMB substrate solution are loaded. The absorbances of the microwell are read at 450 nm.
 
 
 
STORAGE - Keep streptavidin-HRP at 4℃ and protect it from prolonged exposure to light. Aliquot all other reagents and store at -20℃ to -70℃ in a manual defrost freezer.
 
 
 
Plate Preparation
 
 
Dilute the capture antibody to the working concentration in CBS. Immediately coat a 96-well microplate with 100μL per well of the diluted capture antibody. Seal the plate and incubate overnight at 4℃.
 
Aspirate each well and wash with at least 300μl wash buffer, repeating the process two times for a total of three washes. Complete removal of liquid at each step is 3. essential for good performance. After the last wash, remove any remaining wash buffer by inverting the plate and blotting it against clean paper towels.
 
Block plates by adding 300 μL of blocking buffer to each well. Incubate at room temperature for a minimum of 1 hour.
 
Repeat the aspiration/wash as in step 2. The plates are now ready for sample addition.
 
 
 
 
Assay Procedure
 
 
Add 100 μL of sample or standards in sample dilution buffer per well. Seal the plate and incubate 2 hours at room temperature.
 
Repeat the aspiration/wash as in step 2 of plate preparation.
 
Add 100 μL of the detection antibody, diluted in antibody dilution buffer, to each well. Seal the plate and incubate 1 hour at room temperature.
 
Repeat the aspiration/wash as in step 2 of plate preparation.
 
Add 100 μL of Streptavidin-HRP to each well. Incubate for 1 hour at room temperature.
 
Repeat the aspiration/wash as in step 2 of plate preparation.
 
Add 200 μL of substrate solution to each well. Incubate for 20 minutes at room temperature ( if substrate solution is not as requested, the incubation time should be optimized ). Avoid placing the plate in direct light.
 
Add 50 μL of stop solution to each well. Gently tap the plate to ensure thorough mixing.
 
Determine the optical density of each well immediately, using a microplate reader set to 450 nm.
 
 
 
 
Calculation of results
 
 
Calculate the mean absorbance for each set of duplicate standards, controls and samples. Subtract the mean zero standard absorbance from each.
 
Construct a standard curve by plotting the mean absorbance for each standard on the y-axis against the concentration on the x-axis and draw a best fit curve through the points on the graph.
 
To determine the concentration of the unknowns, find the unknowns’ mean absorbance value on the y-axis and draw a horizontal line to the standard curve. At the point of intersection, draw a vertical line to the x-axis and read the concentration. If samples have been diluted, the concentration read from the standard curve must be multiplied by the dilution factor.
 
Alternatively, computer-based curve-fitting statistical software may also be employed to calculate the concentration of the sample.
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 Problems 
 
 
 
 Possible Sources 
 
 
 
 Solutions 
 
 
 
 
 
 No signal 
 
 
 
Incorrect or no Detection Antibody was added
 
 
 
Add appropriate Detection Antibody and continue
 
 
 
 
 
Substrate solution was not added
 
 
 
Add substrate solution and continue
 
 
 
 
 
Incorrect storage condition
 
 
 
Check if the kit is stored at recommended condition and used before expiration date
 
 
 
 
 
 
 
 
 
 
 
 
 
Poor Standard Curve
 
 
 
Standard was incompletely reconstituted or was inappropriately stored
 
 
 
Aliquot reconstituted standard and store at  -70 ℃
 
 
 
 
 
Imprecise / inaccurate pipetting
 
 
 
Check / calibrate pipettes
 
 
 
 
 
Incubations done at inappropriate temperature, timing or agitation
 
 
 
Follow the general ELISA protocol
 
 
 
 
 
Background wells were contaminated
 
 
 
Avoid cross contamination by using the sealer appropriately
 
 
 
 
 
 
 
 
 
 
 
 
 
Poor detection value 
 
 
 
The concentration of antigen in samples was too low
 
 
 
Enriching samples to increase the concentration of antigen
 
 
 
 
 
Samples were ineffective
 
 
 
Check if the samples are stored at cold environment. Detect samples in timely manner
 
 
 
 
 
 
 
 
 
 
 
 
 
High Background 
 
 
Insufficient washes
 
 
Use multichannel pipettes without touching the reagents on the plate
 
 
 
 
 
Increase cycles of washes and soaking time between washes
 
 
 
 
 
TMB Substrate Solution was contaminated
 
 
 
TMB Substrate Solution should be clear and colorless prior to addition to wells
 
 
 
 
 
Materials were contaminated
 
 
 
Use clean plates, tubes and pipettes tips
 
 
 
 
 
 
 
 
 
 
 
 
 
Non-specificity 
 
 
 
Samples were contaminated
 
 
 
Avoid cross contamination of samples
 
 
 
 
 
The concentration of samples was too high
 
 
 
Try higher dilution rate of samples
 
 
 
 
 
 
 
Human TIMP1 ( EPA / CLGI ) ELISA Kit Image
 
 
This standard curve is only for demonstration purposes. A standard curve should be generated for each assay.
 
 
 
Human TIMP1 ( EPA / CLGI ) ELISA Kit Flash
  
This flash is a vivid summary of the assay procedure.
 
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Human TIMP1 / TIMP ( EPA / CLGI ) ELISA Kit Related Products & Topics
 
 
相关研究领域:
 
Enzyme>>Protease & Regulator>>Metalloprotease & Regulator>>Tissue Inhibitor of Metalloproteinase>>TIMP-1/TIMP1
 
Cancer>>Angiogenesis>>Tissue Inhibitor of Metalloproteinase>>TIMP-1/TIMP1
 
Cancer>>Cancer Biomarkers>>TIMP-1/TIMP1
 
蛋白:
 
 
 
 
Molecule
 
Species
 
Description //For Detailed Info.------CLICK!
 
Cat No
 
 
 
TIMP-1/TIMP1
 
Human
 
TIMP-1/TIMP1 Protein, Recombinant  
 
10934-H08H
 
 
 
TIMP-1/TIMP1
 
Human
 
TIMP1/TIMP1 Protein, Recombinant  
 
10934-HNAH
 
 
 
TIMP-1/TIMP1
 
Mouse
 
TIMP-1/TIMP1 Protein, Recombinant  
 
50342-MNAH
 
 
 
 
抗体:
 
 
 
 
Molecule
 
Application
 
Description //For Detailed Info.------CLICK!
 
Cat No
 
 
 
Human TIMP-1/TIMP1
 
ELISA
 
Mouse Monoclonal Antibody
 
10934-MM02
 
 
 
Human TIMP-1/TIMP1
 
WB, ELISA
 
Mouse Monoclonal Antibody
 
10934-MM03
 
 
 
Human TIMP-1/TIMP1
 
ELISA
 
Rabbit Polyclonal Antibody
 
10934-RP01
 
 
 
Human TIMP-1/TIMP1
 
WB
 
Rabbit Polyclonal Antibody Antigen Affinity Purified)
 
10934-RP04
 
 
 
 
 
Background
 
 
Metalloproteinase inhibitor 1, also known as Tissue inhibitor of metalloproteinases 1, Fibroblast collagenase inhibitor, Collagenase inhibitor, CLGI and TIMP1, is a secreted protein which belongs to the protease inhibitor I35 (TIMP) family. TIMP1 complexes with metalloproteinases (such as collagenases) and irreversibly inactivates them by binding to their catalytic zinc cofactor. TIMP1 also mediates erythropoiesis. Unlike IL-3, it is species-specific, stimulating the growth and differentiation of only human and murine erythroid progenitors. TIMP1 is known to act on MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-10, MMP-11, MMP-12, MMP-13 and MMP-16. TIMP1 does not act on MMP-14. Excessive TIMP1 was deleterious to ovulation and embryo development. Novel TIMP1 modulating therapies may be developed to alleviate infertility in women with endometriosis.
Tissue inhibitors of metalloproteinases (TIMP) family are natural inhibitors of the matrix metalloproteinases (MMPs), the zinc enzymes involved in extracellular matrix maintenance and remodeling. The TIMP family encompasses four members ( TIMP1, TIMP2, TIMP3, TIMP4 ), and the inducible form TIMP1 is a glycoprotein expressed by a variety of cell types. It inhibits most MMPs except membrane-type MMP subfamily by forming non-covalent binary complex, but also interacts with the hemopexin-like (PEX) domain of pro MMP-9. TIMP1 has been identified as a multifunctional molecule with divergent functions. It exhibits keratinocyte and fibroblast growth factor-like activity and acts as a cell survival factor, and additionally participates in tumor metastasis, angiogenesis, inflammatory responses, as well as CNS homeostasis. Thus, TIMP1 overexpression may serve to help identify patients with particularly aggressive disease for adjuvant treatments.
 
 
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