FABP3 ELISA Kit

Catalog No. ABIN956147

Product Details FABP3 ELISA Kit

Target: FABP3 (Fatty Acid Binding Protein 3, Muscle and Heart (FABP3))

Reactivity: Mouse

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Application: ELISA

Purpose: Our high sensitivity mouse H-FABP kit is offered as a tool for investigation of heart damage in mouse models of cardiovascular disease.

Sample Type: Plasma, Serum

Analytical Method: Quantitative

Components: Anti-mouse H-FABP antibody coated microtiter plate with 96 wells (provided as 12 detachable strips of 8)
HRP Conjugate Reagent, 11 mL
Mouse H-FABP Calibrator (lyophilized)
10X Diluent (25 mL)
20X Wash Solution (50 mL)
TMB Reagent (One-Step), 11 mL
Stop Solution (1N HCl), 11 mL.

Material not included: Precision pipettes and tips
Distilled or de-ionized water
Polypropylene or glass tubes
Vortex mixer
Absorbent paper or paper towels
Micro-Plate incubator/shaker mixing speed of ~150 rpm
A microtiter plate reader capable of measuring absorbance at 450 nm, with a bandwidth of 10 nm or less and an OD range of 0-4 OD
Graph paper (PC graphing software is optional)

Application Details

Plate: Pre-coated

Protocol: The Mouse H-FABP ELISA is based on a solid phase enzyme-linked immunosorbent assay (ELISA). The assays uses an affinity purified anti-mouse H-FABP antibody for solid phase (microtiter wells) immobilization and a horseradish peroxidase (HRP) conjugated anti-mouse H-FABP antibody for detection. The test sample is diluted and incubated with conjugate in the microtiter wells for 60 minutes. This results in mouse H-FABP molecules being sandwiched between the immobilization and detection antibodies. The wells are then washed to remove unbound HRP labeled antibodies. A solution of TMB Reagent is added and incubated for 20 minutes at room temperature, resulting in the development of a blue color. The color development is stopped with the addition of Stop Solution, and the color is changed to yellow and measured spectrophotometrically at 450 nm. The concentration of H-FABP is proportional to the optical density.

Sample Preparation:

Serum or EDTA plasma may be used in the assay. Avoid use of heparin plasma. Baseline levels of H-FABP are in the range of 1 - 2 ng/mL and can increase to 30 ng/mL or higher in rats following cardiac injury. Similar findings are expected for mice. We recommend that samples be diluted 5-fold prior to assay. This may be achieved by mixing 50 µL of each test sample with 200 µL of 1x diluent.

Assay Procedure:

1. Secure the desired number of coated wells in the holder.
   2. Dispense 100 µL of calibrators and samples into the wells (we recommend that samples be tested in duplicate).
   3. Add 100 µL of enzyme conjugate reagent into each well.
   4. Incubate on an orbital micro-plate shaker at 100-150 rpm at room temperature (18-25°C) for 60 minutes.
   5. Wash and empty the microtiter wells 5 times with 1X wash solution. This may be performed using either a plate washer (400 µL/well) or with a squirt bottle. The entire wash procedure should be performed as quickly as possible.
   6. Strike the wells sharply onto adsorbent paper or paper towels to remove all residual droplets.
   7. Dispense 100 µL of TMB Reagent into each well.
   8. Gently mix on an orbital micro-plate shaker at 100-150 rpm at room temperature (18-25°C) for 20 minutes.
   9. Stop the reaction by adding 100 µL of Stop Solution to each well.
   10. Gently mix. It is important to make sure that all the blue color changes to yellow.
   11. Read the optical density at 450 nm with a microtiter plate reader within 5 minutes.

Calculation of Results:

1. Calculate the average absorbance values (A450) for each set of reference calibrators, and samples.
   2. Construct a calibration curve by plotting the mean absorbance obtained from each reference calibrator against its concentration in ng/mL on linear graph paper, with absorbance values on the vertical or Y-axis and concentration on the horizontal or X-axis.
   3. Using the mean absorbance value for each sample, determine the corresponding concentration of H-FABP in ng/mL from the calibration curve.
   4. Multiply the derived concentration by the dilution factor to determine the actual concentration of H-FABP in the sample.
   5. If available, PC graphing software may be used for the above steps.
   6. If the OD450 values of samples fall outside of the calibration curve, samples should be diluted appropriately and re-tested.

Restrictions: For Research Use only

Handling

Storage: -20 °C

Storage Comment: The reference calibrator provided should be stored frozen at or below -20°C on receipt. The remainder of the kit should be stored at 4°C and the microtiter plate should be kept in a sealed bag with desiccant to minimize exposure to damp air. Test kits will remain stable until the expiration date provided that the components are stored as described above.

Expiry Date: The expiry date is stated on the label.

Target Details for FABP3

Target: FABP3 (Fatty Acid Binding Protein 3, Muscle and Heart (FABP3))

Alternative Name: H-FABP (FABP3 Products)

Synonyms: Fabph-1 ELISA Kit, Fabph-4 ELISA Kit, Fabph1 ELISA Kit, Fabph4 ELISA Kit, H-FABP ELISA Kit, Mdgi ELISA Kit, FABP11 ELISA Kit, M-FABP ELISA Kit, MDGI ELISA Kit, O-FABP ELISA Kit, FABP ELISA Kit, FABP-3 ELISA Kit, fabp3 ELISA Kit, fatty acid binding protein 3 ELISA Kit, fatty acid binding protein 3, muscle and heart ELISA Kit, FABP3 ELISA Kit, Fabp3 ELISA Kit, fabp3 ELISA Kit

Background: Fatty acid-binding proteins are cytoplasmic proteins of about 15 kDa that bind long chain fatty acids and play an important role in fatty acid metabolism. Different types of FABP have been detected including Heart FABP (H-FABP), Liver FABP and Intestinal FABP. Human cardiac muscle has a high content of H-FABP (10-20 mol % of cytoplasmic proteins) and H-FABP is a sensitive biomarker of myocardial necrosis that can be used to confirm or exclude a diagnosis of acute myocardial infarction (AMI). In AMI, H-FABP is rapidly released from damaged cardiomyocytes into the circulation due to its solubility and small size. Human clinical studies indicate that H-FABP levels are significantly elevated above threshold within 3 hours of AMI and subsequently return to normal values in 12 to 24 hours. H-FABP has also been identified as a potential serum biomarker for stroke that is superior to either neuron specific enolase or S100B.

Pathways: Monocarboxylic Acid Catabolic Process