Bovine Serum Albumin (BSA) ELISA Kit

Catalog No. ABIN6579155

Product Details

Target:

Bovine Serum Albumin (BSA)

Reactivity: Cow

Detection Method: Colorimetric

Method Type: Competition ELISA

Detection Range: 0.62 ppm - 300 ppm

Minimum Detection Limit: 0.62 ppm

Application: ELISA

Purpose: The kit is a competitive ELISA intended for the quantitative determination of BSA in dairy foodstuff. This kit is not suitable for the determination of BSA in heat treated foodstuff (the temperature of the heat processing should not exceed 100 °C).

Sample Type: Food

Analytical Method: Quantitative

Specificity: The assay is specific for Bovine serum albumin. Cross-reactivity with Bovine α-casein is 0.3 % , with Bovine β-casein 0.1, with Bovine κ-casein 0.3 % and with β-lactoglobulin 1.4 %. Cross-reactivity with sheep, goat and pig albumin is less than 0.01 %. No interference with Beef meat, Pork meat, Chicken meat, Fish meat, Peanut, Walnut, Hazelnut, Almond, Cashew nut, Pine nut, Coconut, Sunflower seed, Egg white, Egg yolk, Apricot, Wheat, Oat, Barley, Rye, Mustard, Celery, Beans, Pea, Soy, Sesame, Lentil, Chickpea, Rice , Maize.

Components: Microtitre plate with a lid, Calibrators, Control samples, Extraction buffer concentrate, Conjugate concentrate, Conjugate diluent, Wash solution concentrate, TMB, Stop solution, Vial for diluting conjugate

Material not included: In addition to usual laboratory equipment, following items are necessary, - distilled or de-ionized water, - centrifuge with the minimal R.C.F. value of 1,800 g, the volume of tubes must be more than 11 ml, - adjustable micro-pipettes 50, 200 and 1 000 μL, - adjustable repeating dispensers with the volume ranging between 50 - 300 μL, - vortex, - laboratory scale weighing accuracy to hundredths of grams, - grinding mortar or splinter grinder (for treating solid samples), - micro-titre plates reader, equipped with 450 nm filter, - micro-titre plate washer or repeating dispenser for dispensing volume up to 350 μL, - vials or tubes for diluting of extracts.

Application Details

Application Notes: Optimal working dilution should be determined by the investigator.

Sample Volume: 150 μL

Assay Time: 1 h

Protocol: Immuno-enzymatic determination of BSA is based on application of a competition system. In the wells of microtitre plate, walls of which are coated with sheep anti - BSA antibody, the sample to be analysed, while combined with BSA conjugate, is homogenised with horse - radish peroxidase. During the step of subsequent incubation, BSA will be bound to the wells walls. BSA present in the sample and BSA bound in the conjugate will compete mutually for access to the binding sites present in limited number in the antibody against BSA. After subsequent incubation step, the wells are washed and the peroxidase bound to the wells walls is detected by means of a chromogenic substrate (TMB), added to the system. Intensity of thus developed colouration is inversely proportional to BSA concentration in calibrators and analysed samples.

Reagent Preparation:

Working solution of conjugate: Prepare a functional solution of the conjugate by using an 30-fold dilution of the concentrated conjugate. For example, using a pipette, measure out 100 μL of the conjugate concentrate into the conjugate dilution vial and add 3 mL of buffer to dilute the conjugate (pink). This volume will be sufficient for one half of the microtitre plate (48 wells). Make sure to use the functional solution of the conjugate in one day. Washing solution: Prepare the washing solution by a 30-fold dilution of the concentrate. For example: dilute 25 mL with 475 mL of distilled water. This quantity will be sufficient, with a reasonable surplus, to treat the volume of all wells of one microtitre plate (96 wells). The diluted washing solution can be stored at temperatures between 2 and 8 °C until the expiration date of the kit. Preparation of extraction buffer: Dilute the concentrate of extraction buffer 5 times with distilled water. For example, to extract 50 samples you will need (including extra solution) 250 mL of undiluted extraction buffer, to prepare it, blend 50 mL of extraction buffer concentrate with 200 mL of distilled water.

Sample Collection: Food (not thermaly treated, the temperature of the heat processing should not exceed 100 °C)

Sample Preparation:

PREPARATION OF SAMPLES Solid sample: Grind the material in grinding mortar or grind it in splinter grinder to obtain powder material. Loose sample: Use as is, but make sure it's properly homogenised. Samples of different nature: For example, salamis and pates are extracted directly. Keep in mind that these samples tend not to be, as to their content of milk proteins, homogenous and so it's essential to choose the correct methodology of sample extraction. Liquid samples: They are processed without any modification. SAMPLE EXTRACTION Add 5 mL of diluted extraction buffer to 0.5 g of solid sample or 0.5 mL of liquid sample contained in a suitable sealing vial. To facilitate the extraction process, shake the vial for 30 minutes. When the extraction is completed, separate the content of the vial in a centrifuge for 10 minutes at 1,800 g and then extract a sample of the supernatant liquid. STORAGE OF EXTRACTS Keep the supernatants in the frozen state at temperatures below -18 °C.

Assay Procedure:

• Insert 150 μL of the calibrator or the sample + 50 μL of the diluted solution of the conjugate. - Cover the frame with a lid, incubate for 60 minutes at 18 - 25 °C, no shaking. - Suck out and rinse 4 times with the diluted washing solution. - Insert into every well 200 μL of TMB substrate. Incubate for 10 min at 18 - 25 °C in the dark. - Stop the reaction by adding 50 μL of the STOP solution. Measure the colour change at 450 nm.

Calculation of Results:

Read the results off the calibration curve.We recommend to apply the square regression, since other evaluation methods can give slightly different results.

Assay Precision: Intra-assay: 3 samples were tested in 25 replicates, CVs were ≤ 14.7% Inter-assay: 3 samples were tested 10-times, CVs were ≤ 13.6%

Restrictions: For Research Use only

Handling

Preservative: Sodium azide

Precaution of Use: The stop solution contains 2 M hydrochloric acid. Skin or eye contact can cause irritation. In case of staining with stop solution wash the afflicted place with water. Some substances are preserved with sodium azide. Sodium azide may react with lead, copper, or brass, this may create corresponding combustible azides. That is why, when disposing reagents, they have to be flushed with a large amount of water.

Storage: 4 °C

Storage Comment: 2-8°C

Target Details

Target:

Bovine Serum Albumin (BSA)

Alternative Name: Bovine serum albumin (BSA ELISA Kit Abstract)

Synonyms: albumin, ALB

Background: Bovine serum albumin is present in bovine serum (approx. 6 % ) and also in whey fraction of milk.

Molecular Weight: 66.4

Pathways: Lipid Metabolism