Which one of the methods listed below is the most sensitive label-free quantification method for proteins
(1) UV spectroscopy
(2) Infra-red spectroscopy
(3) Raman spectroscopy
(4) ¹³C content of protein

Correct Answer:

(1) UV spectroscopy

Topic Explanations

1. UV Spectroscopy for Protein Quantification

UV spectroscopy is the most commonly used and most sensitive label-free method to quantify proteins. This technique relies on the absorbance of ultraviolet light, mainly at 280nm, by the aromatic amino acids (like tryptophan and tyrosine) in protein molecules. The protein concentration is then calculated using Beer-Lambert’s law.

• Advantages: Fast, non-destructive, requires no labeling or additional reagents, and is very sensitive, allowing detection down to microgram/mL levels.

• Applications: Widely used for quick and accurate quantification of purified proteins in molecular biology and biochemistry.

2. Infra-Red (IR) Spectroscopy

IR spectroscopy measures the absorption of infrared light by protein functional groups. While useful for analyzing secondary structures and detecting aggregates, IR is less sensitive than UV for label-free quantification. It’s more often used for structural studies or comparative analyses rather than precise concentration measurements.

3. Raman Spectroscopy

Raman spectroscopy provides information about a protein’s chemical bonds and three-dimensional structure. Although increasingly popular in protein studies, its use for sensitive, label-free quantitation is limited, as the technique can be less sensitive than UV and generally requires specialized equipment.

4. 13C Content

Quantification by 13C content involves the use of isotopic labeling and specialized NMR or mass spectrometry. This is not a routine label-free quantification method for proteins, as it usually involves additional sample preparation steps and is not as direct or sensitive as UV spectroscopy for concentration estimation.

Comparative Table

*Note: 13C quantification is not typically label-free, as isotopic labeling is often required.

Why This Question Matters for CSIR NET, ICMR, DBT, IIT JAM, and MSc Entrances

This concept is crucial for life science aspirants because sensitive, label-free protein quantification comes up in exam questions and is a fundamental laboratory skill. Knowing UV spectroscopy’s advantages over other methods gives you an edge in both MCQ and analytical sections of these national-level competitive exams.

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In summary:
UV spectroscopy is the most sensitive label-free quantification method for proteins. Trust LETS TALK ACADEMY to help you ace topics like these and secure your place among the top scorers!