Intranuclear staining of proteins in heterogeneous cell populations and verification of nuclear localization by flow cytometric analysis.
We describe techniques to detect nuclear transcription factors in thymocyte subsets using flow cytometry. We have adapted a method that minimizes autofluorescence of fixed cells, thereby allowing the detection of proteins expressed at low levels. An accompanying method in which the cytoplasm is removed from stained cells allows the confirmation of nuclear localization. These methods combine to provide a sensitive alternative approach for detecting nuclear proteins within heterogeneous cell populations.
Doan
Loretta L
LL
Institute for Cellular Therapeutics, University of Louisville Medical School, Louisville, KY 40202, USA.
Tanner
Michael K
MK
Grimes
H Leighton
HL
D000818:Animals; D000906:Antibodies / Q000276:immunology; D002467:Cell Nucleus / Q000276:immunology / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000276:immunology; D051766:Early Growth Response Protein 1; D005434:Flow Cytometry / Q000379:methods*; D017874:Immediate-Early Proteins; D051379:Mice; D009687:Nuclear Proteins / Q000276:immunology; D011518:Proto-Oncogene Proteins / Q000276:immunology; D019253:Proto-Oncogene Proteins c-bcl-2; D013194:Staining and Labeling / Q000379:methods*; D013950:Thymus Gland / Q000166:cytology / Q000276:immunology; D014157:Transcription Factors / Q000276:immunology
D000906:Antibodies; D004268:DNA-Binding Proteins; D051766:Early Growth Response Protein 1; C486109:Egr1 protein, mouse; D017874:Immediate-Early Proteins; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D014157:Transcription Factors; C052617:Bcl2 protein, mouse