Embedding of neural tissue in agarose or glyoxyl agarose for vibratome sectioning.
Agarose was used to embed the brain or spinal cord of lampreys or rats before cutting vibratome sections. Agarose embedding was compatible with immunocytochemistry or the use of horseradish peroxidase as a neuroanatomical tracer. Concentrated agarose with high intrinsic gel strength was optimal for embedding glutaraldehyde fixed neural tissue. A quick procedure was to blot tissue and embed in 5% (w/v) Sigma type I-A or Litex type LSL agarose at 45-55 C dissolved in 50 mM neutral-pH TRIS buffer before cutting 50-100 microns vibratome sections. An alternative procedure that improved retention of tissue sections in the agarose was to rinse the tissue in H2O, blot and embed in 5% (w/v) Sigma type I-A or Litex type LSL agarose at 45-55 C dissolved in H2O, then equilibrate the block overnight in buffer. Phosphate buffer prevented complete dissolving of agarose. Tissue could be covalently linked to the embedding matrix using a novel aldehyde-derived agarose (NuFix, FMC BioProducts). Slices of spinal cord from neonatal rats could be cut after embedding in 5% FMC SeaPrep agarose in rat Ringer's at 23-26 C.
Sallee
C J
CJ
Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri 63110.
Russell
D F
DF
D000818:Animals; D001921:Brain / Q000166:cytology*; D002021:Buffers; D006735:Horseradish Peroxidase; D007798:Lampreys; D008867:Microtomy; D051381:Rats; D012685:Sepharose; D013116:Spinal Cord / Q000166:cytology*; D016610:Tissue Embedding / Q000379:methods*
D002021:Buffers; D012685:Sepharose; D006735:Horseradish Peroxidase