Development of Protein Markers for Quantitative Western Blot Analysis

October 8, 2024 Colloquia Presenters
Department of Biological, Human & Life Sciences

Development of Absolute Quantification Markers for Western Blot

Western blot is one of the most prevalent methods in protein analysis studies. It is also commonly used for quantification studies. However, with current methodology, Western blot is only semi-quantitative at best due to non-linear controls, signal oversaturation, variations in the interpretation of blot image analysis, and other variables. These issues often lead to misinterpretations of data. To measure the absolute amount of protein using Western blot, we are developing a set of standard protein markers. With such markers, Western blot can be performed quantitatively similar to other biological sample measurements, such as ELISA and protein concentration measurement. Using such markers, issues, such as discrepancies in total protein loaded in each sample and at different times, and transfer inconsistency, can be eliminated. The set of markers were purified and tested for quantifying housekeeping proteins such as tubulin and histone h3 proteins in human embryonic kidney 293 cells. The quantitative ladder showed consistent results in creating a standard curve for quantification, with a high R2 value. These results support a reliable and more accurate quantification system for Western Blot.

RESEARCHERS: Vaishnavi Konda, Irvington High School '25; Anh Le, Gunn High School '25; Ayana Modi, Los Altos High School '25; Mark Beliaev, Los Altos High School '26; Amaan Mohammed, Dublin High School '25; Anand George, Junipero Serra High School '25; Eben Ginto, Leigh High School '25; Iris Singh, Design Tech High School '26; Andy Zhang, Mission San Jose High School '27

ADVISOR: Wang Lab Molecular Biology

KEYWORDS: Western Blot | Protein Analysis | Protein Quantification | Molecular Biology