1 Invited Review Cellular Physiol Biochem 2000;10:1-12 Accepted: January 14, 2000 Cellular Physiology Cellular Physiology Cellular Physiology Cellular Physiology Cellular Physiology and Biochemistr and Biochemistr and Biochemistr and Biochemistr and Biochemistry Copyright © 2000 S. Karger AG, Basel Fax +41 61 306 12 34 E-Mail karger@karger.ch www.karger.com © 2000 S. Karger AG, Basel 1015-8987/00/0101-0001$17.50/0 Accessible online at: www.karger.com/journals/net The Use of Xenopus laevis Oocytes for the Fun- ctional Characterization of Heterologously Ex- pressed Membrane Proteins Carsten A. Wagner, Björn Friedrich, Iwan Setiawan, Florian Lang and Stefan Bröer Department of Physiology I, University of Tübingen, Gmelinstr. 5, 72076 Tübingen, Germany Carsten A. Wagner Department of Physiology I, University of Tübingen, Gmelinstr. 5, 72076 Tübingen (Germany) Tel. +49 7071 2973071, Fax +49 7071 293073, E-Mail carsten.wagner@uni-tuebingen.de Abstract The oocytes of the South African clawed frog X. laevis are widely used for the expression of heterologous proteins. The functional characterization of membrane proteins in particular has significantly profited from the use of this expression system. Heterologous cRNA can easily be injected and protein expression and function be studied with several techniques. This review will give a short overview into the variety of methods applicable. They span from different electrophysiolo- gical methods such as two electrode voltage clamp, patch clamp and ion-selective electrodes over cytochemistry to protein biochemistry. In spite of the wide usage of Xenopus oocytes, caution should be taken interpreting the results of protein expression. Heterologous proteins may either interact with endogenous proteins, the background of endogenous protein function may be relatively high, or altered protein behaviour may occur due to differences of the ambient temperature or altered cellular environment. Introduction The South-African clawed frog X. laevis has been kept in captivity and used in laboratory medicine for many years as diagnostic tool for pregnancy tests [1] as laboratory animal for muscle and kidney physiology, developmental biology [1,2] and more recently for the use of its oocytes. Gurdon et al. [3] demonstrated that X. laevis oocytes were able to synthesise haemoglobin upon injection of the corresponding mRNA. Besides their ability to efficiently translate exogenous mRNA into proteins, Xenopus oocytes provide a number of advantages. X. laevis is easily kept and bred in captivity and therefore available from commercial suppliers at low costs [4]. The oocytes are easily prepared and because of their large diameter of 1.1 –1.3 mm easy to handle. Furthermore, as the oocytes are normally deposited into an unfavourable environment, they are resistant and fully equipped with all substrates they need for growth and efficient translation of proteins (Fig. 1). Due to their independence from the necessity to take up nutrients from the environment they express only a small number of endogenous membrane transport systems which gives a low background to the heterologously expressed proteins. Key Words Xenopus laevis • Oocyte physiology • Membrane proteins • Review • Voltage clamp • Immuno- histochemistry