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Flow Cytometry01:23

Flow Cytometry

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The development of flow cytometry techniques began in 1934 with initial attempts by Andrew Moldavan, a bacteriologist who counted the cells in a flowing capillary system. Moldavan pumped cells through a capillary tube focused under a microscope for visualization. The invention of photometry allowed the measurement of differentially-stained cells, and Louis Kamentsky developed the first multiparameter flow cytometer in 1965 to identify and count the cancer cells in cervical tissue specimens.
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Enzymatic Modification and Flow Cytometry Assessment of Yeast Surface Displayed Proteins
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Enzymatic Modification and Flow Cytometry Assessment of Yeast Surface Displayed Proteins

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Surface-decorated S. cerevisiae for flow cytometric array immunoassay.

Wen-Jun Lan1, Yan-Min Lin2, Zheng-Hua Men2

  • 1College of Bioengineering and Shandong Provincial Key Laboratory of Microbial Engineering, Qilu University of Technology, Jinan, Shandong, 250353, China. lanwenjun0522@163.com.

Analytical and Bioanalytical Chemistry
|July 6, 2017
PubMed
Summary
This summary is machine-generated.

This study presents a novel flow cytometric array immunoassay using surface-decorated Saccharomyces cerevisiae (yeast) for protein quantification. This yeast-based method overcomes antibody limitations, offering a sensitive and accurate diagnostic tool for biomarkers like Cytokeratin 19 fragment and neuron-specific enolase.

Keywords:
Adipic dihydrazideFlow cytometric array immunoassayGlutaraldehydeSaccharomyces cerevisiaeSodium periodate

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Quantitative Metabolomics of Saccharomyces Cerevisiae Using Liquid Chromatography Coupled with Tandem Mass Spectrometry
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Area of Science:

  • Biotechnology
  • Analytical Chemistry
  • Immunology

Background:

  • Existing cell-based bio-beads for protein quantification have limitations in antibody selection.
  • Development of novel immunoassay platforms is crucial for sensitive and specific biomarker detection.

Purpose of the Study:

  • To develop a surface-decorated Saccharomyces cerevisiae platform for flow cytometric array immunoassay.
  • To evaluate the performance of this yeast-based immunoassay for quantifying cancer biomarkers Cytokeratin 19 fragment (Cyfra21-1) and neuron-specific enolase (NSE).

Main Methods:

  • Saccharomyces cerevisiae was modified with fluorescein isothiocyanate (FITC) and oxidized to create aldehyde groups for conjugation.
  • Adipic dihydrazide and glutaraldehyde were used for surface functionalization, followed by antibody immobilization.
  • Flow cytometry was employed to validate antibody conjugation and quantify Cyfra21-1 and NSE using phycoerythrin (PE)-labeled secondary antibodies.

Main Results:

  • FITC-labeled yeast generated distinct populations for multiplexing.
  • PE-labeled antibody confirmed successful monoclonal antibody coating on yeast surfaces.
  • The assay achieved a limit of detection (LOD) of at least 0.4 ng/mL for Cyfra21-1 and NSE.
  • High precision (R.S.D. <15%) and accuracy (R.E. 0.9-1.1) were demonstrated.
  • Excellent correlation with electrochemiluminescence immunoassay was observed (R=0.9622 for Cyfra21-1, R=0.9918 for NSE).

Conclusions:

  • Surface-decorated Saccharomyces cerevisiae is a viable platform for flow cytometric array immunoassays.
  • This method offers a sensitive, accurate, and potentially cost-effective alternative for protein quantification.
  • The developed immunoassay shows promise for clinical diagnostics, particularly for cancer biomarkers.