Isolation of cardiomyocyte nuclei from post-mortem tissue.
Identification of cardiomyocyte nuclei has been challenging in tissue sections as most strategies rely only on cytoplasmic marker proteins(1). Rare events in cardiac myocytes such as proliferation and apoptosis require an accurate identification of cardiac myocyte nuclei to analyze cellular renewal in homeostasis and in pathological conditions(2). Here, we provide a method to isolate cardiomyocyte nuclei from post mortem tissue by density sedimentation and immunolabeling with antibodies against pericentriolar material 1 (PCM-1) and subsequent flow cytometry sorting. This strategy allows a high throughput analysis and isolation with the advantage of working equally well on fresh tissue and frozen archival material. This makes it possible to study material already collected in biobanks. This technique is applicable and tested in a wide range of species and suitable for multiple downstream applications such as carbon-14 dating(3), cell-cycle analysis(4), visualization of thymidine analogues (e.g. BrdU and IdU)(4), transcriptome and epigenetic analysis.
Strategic Research Center for Stem Cell Biology and Cell Therapy, University of Lund, Sweden.. Olaf.Bergmann@med.lu.se
Olaf
O
Bergmann
Stefan
S
Jovinge
D000818:Animals; D018797:Cell Cycle Proteins / Q000737:chemistry; D002467:Cell Nucleus / Q000737:chemistry*; D003584:Cytological Techniques / Q000379:methods*; D004247:DNA / Q000737:chemistry; D046169:Dermoscopy / Q000379:methods; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D051379:Mice; D032383:Myocytes, Cardiac / Q000648:ultrastructure*; D013194:Staining and Labeling / Q000379:methods
D018797:Cell Cycle Proteins; C490114:Pcm1 protein, mouse; D004247:DNA