Quantification of protein samples using colorimetric assays

Importance of Colorimetric Protein Quantification

Protein quantification plays a central role in biochemical research, quality control and process development. Traditional A280 measurements can be skewed by buffer components or sample impurities. Colorimetric assays offer an alternative by coupling protein-dependent color changes to absorbance measurements, improving accuracy in complex matrices.

Objectives and Study Overview

This application note presents the implementation of four common colorimetric protein assays on Unchained Labs Lunatic and Little Lunatic systems. The goal is to guide users through assay selection, standard curve generation and automated concentration determination of unknown samples.

Methodology and Instrumentation

Samples are mixed with specific reagents to induce a wavelength-dependent color change proportional to protein concentration. Users select the Protein sample type and choose or define a standard curve on the Lunatic interface or Little Lunatic app. Measured absorbance values are plotted against reference standards to generate a calibration curve using various fitting models. Sample concentrations are then calculated from the fitted equation.

Used Instrumentation

• Unchained Labs Lunatic and Little Lunatic microfluidic protein quantification platforms
• Pierce BCA Protein Assay Kit and Related Reagents
• Bio-Rad Bradford Protein Assay Reagents
• Pierce Modified Lowry Protein Assay Kit
• Pierce 660 nm Protein Assay and G-Biosciences Red660 Assay Reagents

Main Results and Discussion

Automated standard curve fitting provides high reproducibility and visible metrics of fit quality, including coefficient of determination (R2). Six curve models are supported: proportional, linear, point-to-point, quadratic (with or without intercept) and four-parameter logistic. Detergent compatibility testing showed negligible interference up to recommended limits (for example, Tween 80 up to 20 %, Triton X-100 up to 0.01 %, SDS up to 0.5 % and CHAPS up to 20 %), enabling accurate measurements in purification and lysis buffers. Results are displayed graphically with reference and sample points and exported in multiple formats (HTML, XML, TXT, CSV, XLSX, PDF).

Benefits and Practical Applications

• High-throughput, automated quantification reduces manual pipetting errors
• Robust performance in the presence of common detergents and buffer additives
• Flexible reporting and data export support integration with laboratory workflows

Future Trends and Potential Applications

Advances may include expanded assay chemistries for nucleic acids or lipids, cloud-based data management, AI-driven curve optimization and further miniaturization to lower reagent consumption. Real-time monitoring in bioprocessing and seamless LIMS integration represent promising next steps.

Conclusion

The Lunatic family provides a versatile, user-friendly solution for colorimetric protein assays. With multiple standard curve options, comprehensive detergent compatibility and flexible reporting, these platforms streamline protein quantification across research, QA/QC and industrial applications.