Tissue Processing

C57BL/6 mice (Harlan) are maintained at St. Jude Children's Research Hospital animal care facility in accordance with the NIH Guidelines for the care and use of laboratory animals. Mice are kept on a twelve-hour light cycle with food and water ad libitum.

For each block of tissue 16 µm cryo-sections are collected onto 51 SuperFrost Plus slides (Fisher Scientific). A total of 101 sections are collected for the E11 block and P42 coronal block and 151 sections for the E15 block, P7 block and P42 sagittal block. The first section of each block is collected on slide 0. The remaining sections are collected on slides 1-50. Each slide contains sections at approximately 800 µm intervals, i.e. slide 1 contains sections 1, 51, etc. For each set of 51 slides generated, slide 0 and every 10 th slide are stained with cresyl violet to be used as reference slides. The remaining 45 slides are each hybridized with a unique probe. Since each section contains multiple pieces of tissue each probe is hybridized to 55 pieces of tissue representing three planes of section and four developmental ages (see Table below ).

For detailed tissue processing protocols please click on Links below:

Animal husbandry

Collection and Fixation

          E11.5 and E15.5

          P7 and P42

Embedding

          E11.5

          E15.5

          P7

          P42

Cresyl violet staining of Reference slides

Solutions

         Phosphate buffered saline (PBS)

                  - 0.1M PBS

         Avertin

                  - Stock solution

                  - Working solution

        Paraformaldehyde

                  -20% Stock solution

                  - 4% Working solution

       Cresyl Violet Acetate