Bca Protein Assay
Bca Protein Assay - Related Videos
10:50
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Cited by 11 •
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10:56
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04:49
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04:43
β-Lactamase-Based Conductimetric Biosensor Assay for Protein-Protein Interactions
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This video demonstrates a conductimetric biosensor assay to study the interaction of an antibody with a target antigen immobilized on a transducer chip. The biosensor is engineered to have a bifunctional chimeric protein — a β-lactamase enzyme with a functional catalytic site fused to a nanobody for binding to the immobilized target antigen. Upon binding, the catalytic hydrolysis of an antibiotic substrate by the β-lactamase enzyme generates an electrical signal — confirming the...
FAQs
In alkaline conditions, peptide bonds in proteins reduce copper(II) ions to copper(I). Bicinchoninic acid then reacts with the copper(I), producing a purple complex. The amount of this complex determines the measured absorbance near 562 nm, so samples containing more protein produce a stronger signal that can be related to protein concentration.
The purple copper-bicinchoninic acid complex has a measurable absorbance near 562 nm. As protein concentration increases, more copper(I) is generated and more colored complex forms, causing the absorbance to rise. Measuring at this wavelength provides the signal used to compare unknown biological samples with protein standards.
The assay is compatible with many detergents, which makes it useful for biological preparations that contain detergent-based extraction components. This compatibility supports protein measurement in lysates and other experimental samples without limiting the assay solely to purified proteins. The resulting concentration can then guide normalization for downstream biology experiments.
Known protein standards establish the relationship between protein concentration and absorbance. The absorbance of an unknown sample is measured under the same assay conditions and compared with that standard curve. This comparison converts the sample signal into an estimated total protein concentration, allowing different lysates or preparations to be normalized.
A typical measurement uses protein standards alongside the biological samples, exposes them to the assay chemistry under alkaline conditions, and measures the resulting absorbance near 562 nm. The sample readings are then compared with the standard curve. Including standards in the measurement connects the color signal to concentration rather than treating absorbance as an isolated value.
The method is useful for quantifying cell or tissue lysates, purified proteins, and other experimental preparations. Researchers can use the measurements to evaluate protein extraction and normalize sample amounts before downstream procedures. This helps make comparisons more consistent when samples will be analyzed by electrophoresis or immunoblotting.
Before electrophoresis or immunoblotting, researchers can use the measured total protein concentration to adjust samples to comparable protein amounts. This application reduces differences caused by unequal loading and supports more meaningful comparisons among experimental preparations. The assay therefore serves as a preparation and normalization step rather than as the downstream analysis itself.