Novel zinc-based fixative for high quality DNA, RNA and protein analysis.
We have developed a reliable, cost-effective and non-toxic fixative to meet the needs of contemporary molecular pathobiology research, particularly in respect of RNA and DNA integrity. The effects of 25 different fixative recipes on the fixed quality of tissues from C57BL/6 mice were investigated. Results from IHC, PCR, RT-PCR, RNA Agilent Bioanalyser and Real-Time PCR showed that a novel zinc-based fixative (Z7) containing zinc trifluoroacetate, zinc chloride and calcium acetate was significantly better than the standard zinc-based fixative (Z2) and neutral buffered formalin (NBF) for DNA, RNA and protein preservation. DNA sequences up to 2.4 kb in length and RNA fragments up to 361 bp in length were successfully amplified from Z7 fixed tissues, as demonstrated by PCR, RT-PCR and Real-Time PCR. Total protein analysis was achieved using 2-D gel electrophoresis. In addition, nucleic acids and proteins were very stable over a 6-14-month period. This improved, non-toxic and economical tissue fixative could be applied for routine use in pathology laboratories to permit subsequent genomic/proteomic studies.
Lykidis
Dimitrios
D
Department of Histopathology, Division of Investigative Sciences, Imperial College London, Hammersmith Hospital Campus, Ducane Road, London, W12 ONN, UK. dimitrios.lykidis00@ic.ac.uk
Van Noorden
Susan
S
Armstrong
Alan
A
Spencer-Dene
Bradley
B
Li
Jie
J
Zhuang
Zhengping
Z
Stamp
Gordon W H
GW
D000085:Acetates / Q000737:chemistry*; D000818:Animals; D017610:Calcium Compounds / Q000737:chemistry; D002712:Chlorides / Q000737:chemistry*; D004247:DNA / Q000032:analysis / Q000592:standards*; D015180:Electrophoresis, Gel, Two-Dimensional; D005260:Female; D005404:Fixatives / Q000737:chemistry*; D005557:Formaldehyde; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction; D011506:Proteins / Q000032:analysis / Q000592:standards*; D040901:Proteomics; D012313:RNA / Q000032:analysis / Q000592:standards*; D016707:Tissue Fixation / Q000379:methods; D014269:Trifluoroacetic Acid / Q000737:chemistry*; D015032:Zinc / Q000737:chemistry*; D017967:Zinc Compounds / Q000737:chemistry*
D000085:Acetates; D017610:Calcium Compounds; D002712:Chlorides; D005404:Fixatives; D011506:Proteins; D017967:Zinc Compounds; D005557:Formaldehyde; D012313:RNA; C016837:zinc chloride; D004247:DNA; D014269:Trifluoroacetic Acid; D015032:Zinc; C120662:calcium acetate