Chapter 9: Counting proteins in living cells by quantitative fluorescence microscopy with internal standards

Methods in Cell Biology
Jian-Qiu WuThomas D Pollard

Abstract

This chapter describes how a confocal microscope can be treated as a spectrophotometer to measure the absolute number of fluorescent molecules in live cells (Wu and Pollard, 2005).(1) The method provides dynamic range of over three orders of magnitude for counting the number of molecules in a single cell. We present a step-by-step guide to measure concentrations in vivo, explaining many of the practical considerations for using this technique. This chapter is meant as a resource for cell biologists, biochemists, and biophysicists interested in quantifying macromolecules involved in their favorite molecular pathways in live cells.

Citations

Jan 9, 2010·Biochemistry·Diego U Ferreiro, Elizabeth A Komives
Oct 16, 2013·The Journal of Cell Biology·Valerie C CoffmanJian-Qiu Wu
Jul 1, 2010·Molecular Biology of the Cell·Vladimir SirotkinThomas D Pollard
May 28, 2011·Plant Physiology·G Wilma van EsseSacco C de Vries
Aug 10, 2010·PloS One·Yuanzheng Gu, Chen Gu
Mar 19, 2014·The Journal of Cell Biology·Matthew AkamatsuThomas D Pollard
May 7, 2014·The Journal of Cell Biology·Ning WangJian-Qiu Wu
May 3, 2014·Molecular Biology of the Cell·John W GossThomas D Pollard
Jul 17, 2014·ELife·Dani L BodorLars Et Jansen
Dec 24, 2014·Proceedings of the National Academy of Sciences of the United States of America·Geoffrey C RollinsSteve Pressé
Nov 16, 2011·The Journal of Cell Biology·Valerie C CoffmanJian-Qiu Wu
May 26, 2017·Molecular Biology of the Cell·Matthew AkamatsuThomas D Pollard
Jul 1, 2017·Molecular Biology of the Cell·Michael M LacyJulien Berro
Dec 14, 2018·Methods and Applications in Fluorescence·K S GruβmayerD-P Herten
Apr 4, 2020·The Journal of Cell Biology·I-Ju LeeDavid Pellman
Jun 21, 2019·Biotechnology Journal·Joshur AngSteve Oh
Jul 3, 2016·Cold Spring Harbor Protocols·Iain M Hagan, Steven Bagley
Feb 2, 2018·Proceedings of the National Academy of Sciences of the United States of America·Caroline Laplante
Dec 5, 2018·Nature Communications·Syed Mukhtar AhmedIan G Macara
Jan 17, 2020·The Journal of Chemical Physics·Jon GarryAndrew D Rutenberg
Sep 2, 2019·Journal of Hazardous Materials·Amanda Pereira da Costa AraújoGuilherme Malafaia
Jan 27, 2021·The Journal of Cell Biology·Kimberly Bellingham-JohnstunCaroline Laplante
May 4, 2010·Neuron·Justin W Taraska, William N Zagotta

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