April 29, 2025

From Cells to Molecules: Western blot assays to investigate cellular
function

Instructor: Claudia Fallini, PhD, URI

Location: URI   
Session 1: July 9 – July 11 ( Location TBD)
Session 2: July 16 – July 18 ( Location TBD)

Course Overview

This module will train students in standard cellular and biochemical techniques aimed at investigating normal or pathological cellular processes. Through a combination of theory and hands-on work, students will learn how to grow and transfect cells, extract proteins, process samples for western blot assays, and quantify protein levels. Students will also learn how to ensure data integrity and proper handling. Some knowledge of basic lab skills (e.g. use of micropipettes, making solutions) is beneficial but not required. The module will be completed
over the course of 2.5 days. Participants who complete the module will receive a certificate of completion.

Learning Outcomes

At the end of this module, students will be able to:

  • Understand the importance and limitations of cell-based methods to study protein
    function and biochemical/molecular processes
  • Familiarize with basic cell culture and transfection methods
  • Learn to process cells to extract and quantify protein fractions
  • Learn how to separate and visualize proteins using SDS-gel electrophoresis
  • Analyze and compare protein levels between samples using proprer controls

Lab Report

Students will be expected to maintain detailed laboratory notes to include:

  • Protocols for cell plating and transfection
  • Quantification of protein concentrations from whole cell lysates
  • Protocols and procedures for gel loading and electrophoresis
  • Membrane preparation and antibody hybridization
  • Results of data quantification (protein levels in different conditions)

Resources Used in this Module:

  • Slide Decks for Lectures
  • Lab Handouts
  • Various Lab Supplies and Equipment

Timeline 

Day 1Day 2Day 3
9:00 AM - 10:00 AMModule Overview and Basics of aseptic technique (lecture)Western blot: theory and practice part 1 (lecture)Secondary antibody hybridization
10:00 AM - 11:00 AMCells splitting, counting, and plating (hands-on)Quantify protein concentration using a plate reader (hands-on)Western blot: theory and practice part 2 (Lecture)
11:00 AM - 12:00 PMCells splitting, counting, and plating (hands-on)Prepare protein samples and load an SDS gel (hands-on)Membrane imaging and data quantification
12:00 PM - 1:00 PMBreakBreak (gels will run during break)
1:00 PM - 2:00 PMCell transfection (lecture and hands-on)Set up of gel transfer (lecture and hands-on)
2:00 PM - 3:00 PMQuantifying transfection effeciency and protein extraction (hands-on)Membrane stain and blocking (hands-on)
3:00 PM - 4:00 PMQuantifying transfection effeciency and protein extraction (hands-on)Primary antibody hybridization (O/N)