BCA Instructions for Protein Content Determination Kit

Micro method 100T/96S

Before formal testing, it is necessary to take 2-3 Make predictions based on samples with significant expected differencesMeasurement significance:

The soluble protein content of the sample is often used for enzyme activity calculation. In addition, soluble protein content is also used for quality analysis of food and other products.

Measurement principle:

Under alkaline conditions, cysteine, cysteine, tryptophan, tyrosine, and peptide bonds in proteins can Cu²⁺restoreCu⁺；2 moleculeofBCAandCu⁺Combining to generate a purple complex, in 540-595nmThere is an absorption peak,562nmThe absorption peak is strongest at this point.

BCA protein content determination kit is readily available in stock

composition

Preparation of working fluid:Before use, please refer to the volume of the proposed working fluid (sample size)×0.2ml）, transfer the reagentsA andB according to 50：1 Mix the proportions, coverMix thoroughly after tightening.

The specific product parameters are subject to the parameters in the product manual received

Self provided instruments and supplies:

Desktop centrifuge, constant temperature water bath, visible spectrophotometer/ELISA reader, trace quartz colorimetric dish/96 Orifice plate, pipette, and distilled water.

Extraction of soluble proteins from samples:

1Liquid samples:Clear liquid samples can be directly measured.

2Organize samplesAccording to organizational quality（g）Volume of extraction solution(ml)for 1：5~10 The proportion of（Suggest weighing the agreement 0.1g Organize, join 1mlExtraction solution (self prepared, choose enzyme extraction buffer, distilled water or physiological saline as needed)Ice bath homogenate,10000rpm，4℃centrifugation 10minTake the supernatant, which is the test solution.

3 Bacteria and cells: according to the number of cells（104 A)Volume of extraction solution（ml）for 500~1000：1 The proportion of（suggestion 500 Ten thousand cellsjoin 1ml extract solution）Ice bath ultrasonic disruption of cells（power 300w, Ultrasound 3 Seconds, intervals 7 Second, total time 3min）And then 10000rpm， 4℃, centrifugation 10minTake the supernatant and place it on ice for testing.

Measurement operation:

1. Visible spectrophotometer/Preheating of ELISA reader 30minAdjust the wavelength to 562 nmAdjust distilled water to zero.

2. Working fluid is placed in 60Preheating in a ℃ water bath30 min.

Notes:Blank tubes and standard tubes only need to be done once.

Calculation formula:

Cpr (mg/ml)=C reference standard×（A Measurement tube -A Blank tube)÷（A Standard tube -A Blank tube）

=0.5×（A Measurement tube -A Blank tube)÷（A Standard tube -A Blank tube）

Cpr (mg/g fresh weight)=C reference standard×（A Measurement tube -A Blank tube)÷（A Standard tube -A Blank tube)÷W

=0.5×（A Measurement tube -A Blank tube)÷（A Standard tube -A Blank tube)÷W W: Sample quality,g

Notes:

BCA Protein content determination kit, suitable for measuring protein concentration in 20-5000μg/ml Sample. Take before measurement 1-2 Conduct a preliminary experiment on a sample, ifA Measurement tube -A Blank tube>1.5The sample needs to be diluted with extraction solution before measurement to ensure the accuracy of the measurement.

BCA protein content determination kit is readily available in stock