Biomedica : revista del Instituto Nacional de Salud · 2005
[Production of polyclonal antibodies to protein iduronate-2-sulphate sulphatase (IDS) and development of a detection system for human recombinant IDS].
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Peña Olga, Sosa Angela, Echeverri Olga, Sáenz Homero, Barrera Luis A
Аннотация
INTRODUCTION: Hunter syndrome is a lysosomal disorder characterized by iduronate 2-sulphate sulphatase (IDS) genetic deficiency. Although MPS type II (Hunter) has no cure, enzyme replacement therapy (ERT) and gene therapy are potential new approaches to treatment.
OBJECTIVE: The Institute for the Study of Innate Errors of Metabolism is currently developing a sulphatase expression system through human recombinant IDS (hrIDS) in Pichia pastoris and Escherichia coli.
MATERIAL AND METHODS: A monitoring method for IDS was developed by production of polyclonal antibodies from rabbit against human IDS. Two New Zealand white rabbits were immunized with commercial IDS. These antibodies were used in a dot-blot method for detection and partial quantification of human recombinant IDS. P. pastoris and E. coli fermentation crude extracts were processed to determine IDS presence.
RESULTS: The antibodies were demonstrably IDS specific, without cross reaction with crude extract contaminant proteins.
CONCLUSION: Therefore, these antibodies can be used to establish an ELISA sandwich system as a method for detecting and quantifying a protein of interest. The antibodies can also be employed in an affinity chromatography step during the IDS purification process.
OBJECTIVE: The Institute for the Study of Innate Errors of Metabolism is currently developing a sulphatase expression system through human recombinant IDS (hrIDS) in Pichia pastoris and Escherichia coli.
MATERIAL AND METHODS: A monitoring method for IDS was developed by production of polyclonal antibodies from rabbit against human IDS. Two New Zealand white rabbits were immunized with commercial IDS. These antibodies were used in a dot-blot method for detection and partial quantification of human recombinant IDS. P. pastoris and E. coli fermentation crude extracts were processed to determine IDS presence.
RESULTS: The antibodies were demonstrably IDS specific, without cross reaction with crude extract contaminant proteins.
CONCLUSION: Therefore, these antibodies can be used to establish an ELISA sandwich system as a method for detecting and quantifying a protein of interest. The antibodies can also be employed in an affinity chromatography step during the IDS purification process.