SERPINA1 ELISA Kit

Catalog No. ABIN956075

Product Details SERPINA1 ELISA Kit

Target: SERPINA1 (serpin Peptidase Inhibitor, Clade A (Alpha-1 Antiproteinase, Antitrypsin), Member 1 (SERPINA1))

Reactivity: Mouse

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Application: ELISA

Purpose: The Mouse Alpha 1-Anti-trypsin ELISA is a highly sensitive two-site enzyme-linked immunoassay (ELISA) for the quantitative determination of Alpha 1-Anti-trypsin (A1AT) in mouse biological samples.

Analytical Method: Quantitative

Characteristics: In this assay the A1AT present in the sample reacts with the anti-A1AT antibody, which has been adsorbed to the surface of polystyrene microtiter wells. After the removal of unbound proteins by washing, anti-A1AT antibody conjugated with horseradish peroxidase (HRP) is added. This HRP-conjugated antibody forms a complex with the previously bound A1AT. Following another washing step, the enzyme bound to the immunosorbent is assayed by the addition of a chromogenic substrate, 3,3',5,5'- tetramethylbenzidine (TMB). The quantity of bound enzyme is proportional to the concentration of A1AT in the sample tested, thus, the absorbance, at 450 nm, is a measure of the concentration of A1AT in the test sample. The quantity of A1AT in the test sample can be interpolated from the calibration curve constructed from the calibrators and corrected for sample dilution.

Components: 1. Diluent Concentrate: One bottle containing 50 mL of a 5X concentrated diluent running buffer.
2. Wash Solution Concentrate: One bottle containing 50 mL of a 20X concentrated wash solution.
3. Enzyme-Antibody Conjugate Concentrate: One vial containing 150 µL of a 100X concentrated affinity-purified anti-mouse A1AT antibody conjugated with HRP in stabilizing buffer.
4. TMB Substrate Solution: One vial containing 12 mL of TMB and hydrogen peroxide in citric acid buffer at pH 3.3.
5. Stop Solution: One vial containing 12 mL of 0.3 M sulfuric acid. WARNING: Avoid contact with skin.
6. Microtiter Plate: Twelve removable eight-well strips in well holder frame. Wells are coated with affinity-purified anti-mouse A1AT.
7. Mouse A1AT Calibrator: One vial containing a lyophilized Mouse A1AT Calibrator.
8. Positive Control: One vial containing 50 µL of serum with 0.1% sodium azide. See the Control Certificate for the concentration.

Material not included: Precision pipettes (2 µL to 200 µL) for making and dispensing dilutions
Test tubes
Microplate washer/aspirator
Distilled or de-ionized H2O Microplate reader
Assorted glassware for the preparation of reagents and buffer solutions
Timer
Vortex mixer

Application Details

Plate: Pre-coated

Reagent Preparation:

1. Diluent Concentrate: The Diluent solution supplied is a 5X concentrate and must be diluted 1:5 with distilled or de-ionized water.
   2. Wash Solution Concentrate: The Wash Solution supplied is a 20X concentrate and must be diluted 1:20 with distilled or de-ionized water. Crystal formation in the concentrate is not uncommon when storage temperatures are low. Warming of the concentrate to 30-35°C before dilution can dissolve crystals.
   3. Enzyme-Antibody Conjugate Concentrate: Calculate the required amount of working conjugate solution for each microtiter plate test strip by adding 10 µL Enzyme-Antibody Conjugate to 990 µL of 1X Diluent for each test strip to be used for testing. Mix uniformly, but gently. Avoid foaming.
   4. TMB Substrate Solution: Ready to use as supplied.
   5. Stop Solution: Ready to use as supplied.
   6. Microtiter Plate: Ready to use as supplied. Unseal Microtiter Pouch and remove plate from pouch. Remove all strips and wells that will not be used in the assay and place back in pouch and re-seal along with desiccant.
   7. Mouse A1AT Calibrator: Add 1.0 mL of distilled or de-ionized water to the lyophilized Mouse A1AT Calibrator and mix gently until dissolved. The calibrator is now at a concentration of 4.40 µg/mL (the reconstituted calibrator should be aliquoted and frozen if future use is intended). Mouse A1AT Calibrators need to be prepared immediately prior to use. Mix well between each step. Avoid foaming.
   8. Positive Control: The concentration and recommended dilution are provided on the Control Certificate. Before use, briefly centrifuge the Positive Control to allow all of the liquid to collect in the bottom of the vial.

Calculation of Results:

1. Subtract the average background value from the test values for each sample.
   2. Using the results observed for the calibrators construct a calibration curve. The appropriate curve fit is that of a four- parameter logistics curve, although a second order polynomial (quadratic) or other curve fits may also be used.
   3. Interpolate test sample values from calibration curve. Correct for sample dilution factor to arrive at A1AT concentration in original sample.
   
   1. Reliable and reproducible results will be obtained when the assay procedure is carried out with a complete understanding of the information contained in the package insert instructions and with adherence to good laboratory practice.
   2. Factors that might affect the performance of the assay include proper instrument function, cleanliness of glassware, quality of distilled or de-ionized water, and accuracy of reagent and sample pipettings, washing technique, incubation time or temperature.

Restrictions: For Research Use only

Handling

Storage: 4 °C

Storage Comment: 1. Complete Kit: The expiration date for the kit is stated on the outer label. The recommended storage temperature is 4°C. Note: See long term storage recommendations below for the Mouse A1AT Calibrator and Positive Control.
2. Diluent: The 5X Diluent Concentrate is stable until the expiration date. The 1X working solution is stable for at least one week from the date of preparation. Both solutions should be stored at 4°C.
3. Wash Solution: The 20X Wash Solution Concentrate is stable until the expiration date. The 1X working solution is stable for at least one week from the date of preparation. Both solutions can be stored at room temperature (RT, 16-25°C) or at 4°C.
4. Enzyme-Antibody Conjugate: Undiluted anti-A1AT-HRP conjugate should be stored at 4°C and diluted immediately prior to use. The working conjugate solution is stable for up to 8 hours.
5. TMB Substrate Solution: The TMB Substrate Solution should be stored at 4°C and is stable until the expiration date.
6. Stop Solution: The Stop Solution should be stored at 4°C and is stable until the expiration date.
7. Microtiter Plate: Anti-mouse A1AT coated wells are stable until the expiration date, and should be stored at 4°C in the sealed foil pouch with desiccant pack.
8. Mouse A1AT Calibrator: The lyophilized Mouse A1AT Calibrator should be stored at 4°C or frozen until reconstituted. The reconstituted calibrator should be aliquoted and stored frozen. Avoid multiple freeze/thaw cycles. The working calibrator solutions should be prepared immediately prior to use and are stable for up to 8 hours.
9. Positive Control: For storage longer than 7 days keep frozen until the expiration date. Storage less than 7 days can be at 4°C. Avoid multiple freeze/thaw cycles.

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

Target Details for SERPINA1

Target: SERPINA1 (serpin Peptidase Inhibitor, Clade A (Alpha-1 Antiproteinase, Antitrypsin), Member 1 (SERPINA1))

Alternative Name: alpha-1 Anti-Trypsin (SERPINA1 Products)

Synonyms: AACT ELISA Kit, ACT ELISA Kit, GIG25 ELISA Kit, A1AT ELISA Kit, PI ELISA Kit, alpha-1-PI ELISA Kit, A1A ELISA Kit, AAT ELISA Kit, PI1 ELISA Kit, PRO2275 ELISA Kit, alpha1AT ELISA Kit, Pi ELISA Kit, Spi1 ELISA Kit, MGC64429 ELISA Kit, serpina1d ELISA Kit, Aps ELISA Kit, SERPINA3-5 ELISA Kit, serpin family A member 3 ELISA Kit, alpha-1-antiproteinase F precursor ELISA Kit, serpin family A member 1 ELISA Kit, alpha-1-antitrypsin ELISA Kit, calumenin ELISA Kit, serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 S homeolog ELISA Kit, serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 ELISA Kit, serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 5 ELISA Kit, SH2B adaptor protein 2 ELISA Kit, serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 3 ELISA Kit, serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 L homeolog ELISA Kit, SERPINA3 ELISA Kit, LOC100008973 ELISA Kit, SERPINA1 ELISA Kit, LOC100328621 ELISA Kit, CALU ELISA Kit, Serpina1 ELISA Kit, serpina1.S ELISA Kit, serpina1 ELISA Kit, SPIA5 ELISA Kit, Sh2b2 ELISA Kit, Calu ELISA Kit, serpina1.L ELISA Kit

Background: Alpha 1-Anti-trypsin is a 52 kDa serine protease inhibitor (serpin) in blood, which protects tissue from enzymes from inflammatory cells, especially elastase. In certain acute phase inflammatory reactions, A1AT is elevated in order to limit the damage caused by activated neutrophil granulocytes and their enzyme elastase. Disorders of A1AT include A1AT deficiency, a hereditary disorder that can lead to severe tissue breakdown during inflammation. This may result in pulmonary emphysema and liver cirrhosis, in severe cases. Genetic variants of A1AT do occur.