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Simultaneous quantification of c-myc oncoprotein, total cellular protein, and DNA content using multiparameter flow
H H Engelhard1, J L Krupka, K D Bauer
1Department of Pathology, Northwestern University Medical School, Chicago, Illinois 60611.
Cytometry
|January 1, 1991
Summary
This study introduces a novel flow cytometry method to simultaneously measure specific proteins, total protein, and DNA. This technique accurately quantifies c-myc oncoprotein levels and cellular protein content, aiding research on cell growth regulation.
Area of Science:
- Cell Biology
- Biochemistry
- Immunology
Background:
- Cellular protein variations can complicate the analysis of specific protein changes.
- Accurate quantification of specific proteins alongside total protein and DNA is crucial for understanding cellular processes.
Purpose of the Study:
- To develop and validate a simultaneous multi-parameter flow cytometry technique.
- To quantify specific cellular proteins (c-myc oncoprotein), total cellular protein, and DNA content concurrently.
Main Methods:
- Utilized dual-laser excitation (UV and 488 nm) and three fluorescent dyes (FITC, SR101, DAPI).
- Employed FITC-labeled antibodies for c-myc oncoprotein, SR101 for total protein, and DAPI for DNA.
- Validated flow cytometry measurements against biochemical methods (densitometry, Lowry technique).
Main Results:
- Flow cytometry measurements showed strong correlation with established biochemical methods.
- Cycloheximide treatment of HL-60 cells led to a rapid decrease in c-myc oncoprotein.
- Observed changes in cell cycle distribution (G0/G1 increase) and a modest decrease in total protein post-treatment.
Conclusions:
- The developed technique offers a rapid and quantitative approach for analyzing cellular growth balance.
- This method is valuable for studying cell differentiation, neoplastic transformation, and responses to radiation or drugs.
