Purification of Anti-Mycobacterium tuberculosis MPT64 Immunoglobulin-Y from Egg-Yolk Supernatant Using Thiophilic Adsorption Chromatography

Authors

  • Sri Agung Fitri Kusuma
  • Muhammad Fadhlillah
  • Intan Timur Maisyarah
  • Shintani Ayunda Khairunnisa
  • Muhammad Yusuf
  • Yaya Rukayadi
  • Toto Subroto

DOI:

https://doi.org/10.36877/pmmb.a0000400

Abstract

The significance of immunoglobulin Y (IgY) as a particular antibody equal to mammalian IgG is well understood. However, due to a lack of reliable purification procedures, producing highly pure IgY remains problematic. In this study, we aimed to optimize the recovery of pure IgY anti-MPT64 using thiophilic adsorption chromatography. The purification of IgY anti-MPT64 was achieved by initial PEG lipid precipitation, then an optimized purification by varying the gradient concentration of elution buffer into five steps gradient (0-20, 20-40, 40-60, 60-80, and 80-100%v/v) for three injection column volume (CV) each and two steps gradient (0-50 and 50-100%v/v) in eight CV for each concentration. The obtained IgY was characterized by SDS-PAGE and dot-blot then determined its content levels using the Lowry method. The results showed that the five steps gradient purification was found to provide a better purity level of IgY than the two steps gradient. However, the IgY content obtained in the two steps gradient purification (2.2632± 0.011 mg/mL) was higher than the five steps gradient purification (1.35482 ± 0.023 mg/mL). Nevertheless, both purified IgY results can recognize MPT64 protein through a dot blot test. Therefore, it can be summarized that thiophilic adsorption chromatography with five steps gradient of purification was an efficient process to obtain a higher purity of IgY anti-MPT64, especially to be targeted as a diagnostic kit component for MPT64 detection.

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Published

2024-03-19

How to Cite

Kusuma, S. A. F., Fadhlillah, M. ., Maisyarah, I. T. ., Khairunnisa, S. A. ., Yusuf, M., Rukayadi, Y., & Subroto, T. (2024). Purification of Anti-Mycobacterium tuberculosis MPT64 Immunoglobulin-Y from Egg-Yolk Supernatant Using Thiophilic Adsorption Chromatography . Progress In Microbes & Molecular Biology, 7(1). https://doi.org/10.36877/pmmb.a0000400

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Section

Original Research Articles
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