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Amplite® Colorimetric BCA Protein Quantitation Assay Kit

. BSA dose responses were measured with Amplite® Colorimetric BCA Protein Quantitation Assay Kit using a clear bottom 96-well plate.
. BSA dose responses were measured with Amplite® Colorimetric BCA Protein Quantitation Assay Kit using a clear bottom 96-well plate.
Ordering information
Price ()
Catalog Number11115
Unit Size
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Additional ordering information
Telephone1-408-733-1055
Fax1-408-733-1304
Emailsales@aatbio.com
InternationalSee distributors
ShippingStandard overnight for United States, inquire for international
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
UNSPSC12171501

OverviewpdfSDSpdfProtocol


The traditional BCA protein assay is widely used for quantifying proteins. However, it is slow and tedious, e.g., you must either heat the BCA reaction at 37 °C for 30 minutes or wait for two hours at room temperature. Amplite®™ Colorimetric BCA Protein Quantitation Assay Kit is a two-component and detergent-compatible assay to determine total protein concentrations. The assay is based on the same copper-chelating reaction and provides comparable accuracy to the traditional BCA protein assay that is either run at high temperature or with longer incubation time. The protein signal is monitored around 560 nm and the assay is completed within 30 minutes. It is convenient, rapid, and robust without high temperature required. Amplite®™ Colorimetric BCA Protein Quantitation Assay Kit can be performed in a convenient 96-well microtiter-plate format and easily adapted to automation with no separation steps required.

Platform


Absorbance microplate reader

Absorbance562 nm
Recommended plateClear bottom

Components


Component A: BCA Solution A50 mL
Component B: BCA Solution B1 mL
Component C: BSA Standard (1 mg/mL)1 mL

Example protocol


AT A GLANCE

Protocol summary
  1. Prepare BCA working solution (50 µL)
  2. Add BSA standards or test samples (50 µL)
  3. Incubate at room temperature for 20 - 60 minutes
  4. Read absorbance at 562 nm (in the range of 540-590 nm) 

Important
Thaw all the kit components at room temperature before use.

PREPARATION OF STANDARD SOLUTION

For convenience, use the Serial Dilution Planner:
https://www.aatbio.com/tools/serial-dilution/11115


BSA Standard
Add 80 µL of 2 mg/mL BSA Standard (Component C) to 320 µL of PBS (not provided) to generate 400 µg/mL BSA standard solution (BS1). Then use 1:2 serial dilutions in PBS to get serially diluted BSA standards (BS2 - BS7). Note: It is necessary to create a standard curve during each assay.

PREPARATION OF WORKING SOLUTION

BCA working solution
  1. Prepare the amount of BCA working solution needed by mixing 50 parts of BCA Solution A (Component A) with 1 part of BCA Solution B (Component B) (50:1, v/v ratio of Solution A: B).
  2. Mix well until the BCA working solution is a uniform, light green color.
    Note     1 mL BCA working solution is enough for 20 tests. 

SAMPLE EXPERIMENTAL PROTOCOL

Table 1.Layout of BSA standards and test samples in a clear bottom 96-well microplate. BS= BSA Standards (BS1 - BS7, 400 to 6.25 µg/mL); BL=Blank Control; TS=Test Samples
BS1 BS1 TS TS
BS2 BS2 ... ...
BS3 BS3    
BS4 BS4    
BS5 BS5    
BS6 BS6    
BS7 BS7    
BL BL    
Table 2.Reagent composition for each well.
Well Volume Reagent
BS1-BS7 50 µL Serial dilutions (400-6.25 µg/mL)
BL 50 µL PBS
TS 50 µL Test samples
  1. Prepare BSA standards (BS), blank controls (BL), and test samples (TS) according to the layout provided in Tables 1 and 2.
  2. Add 50 µL of BCA working solution to each well of BSA standard, blank control, and test samples to make the total assay volume of 100 µL/well.
  3. Incubate the reaction at room temperature for 20 to 60 minutes.
  4. Monitor the absorbance with an absorbance microplate reader at OD 562 nm (in the range of 540 to 590 nm). 

References


View all 15 references: Citation Explorer
Specialized Cell-Free DNA Blood Collection Tubes Can Be Repurposed for Extracellular Vesicle Isolation: A Pilot Study.
Authors: Heatlie, Jessica and Chang, Vanessa and Fitzgerald, Sandra and Nursalim, Yohanes and Parker, Kate and Lawrence, Ben and Print, Cristin G and Blenkiron, Cherie
Journal: Biopreservation and biobanking (2020)
Proteomic Approach for Extracting Cytoplasmic Proteins from Streptococcus sanguinis using Mass Spectrometry.
Authors: El-Rami, Fadi and Nelson, Kristina and Xu, Ping
Journal: Journal of molecular biology research (2017): 50-57
Protein Quantitation and Analysis of Purity.
Authors: Campion, Eva M and Loughran, Sinéad T and Walls, Dermot
Journal: Methods in molecular biology (Clifton, N.J.) (2017): 225-255
[iTRAQ technology combined with 2D-LC-MS/MS to analyze effect of Coptidis Rhizoma on cytochrome P450 isoenzyme expression].
Authors: Yang, Xin and Wang, Qiu-Hong and Wang, Meng and Wang, Yue and Yang, Bing-You and Kuang, Hai-Xue
Journal: Zhongguo Zhong yao za zhi = Zhongguo zhongyao zazhi = China journal of Chinese materia medica (2016): 731-736
Total protein quantitation using the bicinchoninic acid assay and gradient elution moving boundary electrophoresis.
Authors: Kralj, Jason G and Munson, Matthew S and Ross, David
Journal: Electrophoresis (2014): 1887-92
Quantitative evaluation of proteins with bicinchoninic acid (BCA): resonance Raman and surface-enhanced resonance Raman scattering-based methods.
Authors: Chen, Lei and Yu, Zhi and Lee, Youngju and Wang, Xu and Zhao, Bing and Jung, Young Mee
Journal: The Analyst (2012): 5834-8
Quantitation of protein.
Authors: Noble, James E and Bailey, Marc J A
Journal: Methods in enzymology (2009): 73-95
Interferences of suspended clay fraction in protein quantitation by several determination methods.
Authors: Lozzi, I and Pucci, A and Pantani, O L and D'Acqui, L P and Calamai, L
Journal: Analytical biochemistry (2008): 108-14
A comparison of protein quantitation assays for biopharmaceutical applications.
Authors: Noble, J E and Knight, A E and Reason, A J and Di Matola, A and Bailey, M J A
Journal: Molecular biotechnology (2007): 99-111
Biochemical characterization of recombinant mouse amelogenins: protein quantitation, proton absorption, and relative affinity for enamel crystals.
Authors: Ryu, O H and Hu, C C and Simmer, J P
Journal: Connective tissue research (1998): 207-14; discussion 241-6