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Centro de Investigaciones Regionales Dr. Hideyo Noguchi, Universidad Autónoma de Yucatán

2007, Number 3

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Rev Biomed 2007; 18 (3)

Expression of single chain antibody fragments (ScFv) against Helicobacter pylori in the yeast Pichia pastoris

Alonso-Ochoa MD, Zavala-Tapia O, Pedroza-Roldán C

Language: Spanish
References: 32
Page: 168-174
PDF size: 274.44 Kb.


ABSTRACT

Introduction. Helicobacter pylori is one of the most important causal agents of gastric diseases around the world, the opportune detection of the bacterium would avoid the development of gastritis and in some cases gastric cancer. The use of specific antibodies against H. pylori allows the detection of this pathogen in the microbial population of samples; unfortunately the expression of antibodies in some bacterial vectors reduces the efficient expression of high amount of functional protein. The yeast Pichia pastoris allows the expression of great amounts of recombinant protein with high quality; this system can be used for the expression of single chain antibodies against H. pylori for the development of immunodiagnostics.
Material and methods. Identification of reactive clones against H. pylori strains ATCC 49396, N2 and J99 isolated from a ScFv library, characterization of selected clones and isolation of the ScFv gene coding for the fragment of antibody, and construction of the ScFv in the expression vector Pichia pastoris. Induction and quantification of expression of the antibody.
Results and discussion. Ninety six clones were isolated and only 10 clones with the best reactivity and specificity against H. pylori were selected. From these 10 clones we isolate the ScFv from the clone 9D to express its ScFv in the yeast expression vector. The quantification tests showed high levels of protein expression (50 µg/ml) of the recombinant protein at the 72 hours of induction.


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