Since 'nobody' had an answer yet I'd like to encourage you to tell a bit more about your project...e.g.:

- brand / type of the tissue processor you use in your Histology lab

- define age of "mouse / mice pup(s)"

- which 'quality' of tissue preservation is expected, i. e. which part of the animals' body deserves most attention to be an equivalent image after fixation -dehydration - embedding - cutting etc....?

- what is the purpose your processed 'pups' shall serve - guessing you want to embed whole bodies into (paraffin) wax and not resins....

Note added (don't want to teach you!): eventually you can choose from some literature of your predecessors as well as actual researchers/research institutions in using Google as a really strong search data base: using keywords: (without quotation marks):

"Tissue processor processing whole mouse or mice pups for Histology Electron Microscopy"

>>"Tissue processor processing whole mouse or mice pups for Histology"" fixation processing whole mouse pups protocol < or

search phrase:

>" whole mouse tissue processing or mice pups for Histology "< found also

UCSF

Chuang Lab Protocol Updated: 6/27/2001

Created by Pao Tien Chuang and Wanhui Zhang

Paraffin embedding and sectioning of mouse embryos

(valid only for mouse embryos, therefore I am sure that simple interpolation of their processing times for the bigger dimensions of "pups" is / are not a choice):

http://www.cvri.ucsf.edu/~chuang/pdf/Paraffin%20embedding%20and%20sectioning%20of%20mouse%20embryos.pdf

Also, these authors didn't state details on the fixation process prior to embedding...(admitting that - in older/former times - I witnessed "perfusion fixation" as well as filling vessels for microcorrosion casts" of rat and mice pups (e.g. PN-day 1-36).

Nevertheless - hoping someone more competent would chime in as soon as possible - I wish you good luck with your challenging lab-work.

And - last but not least - perhaps you'll let us know how it went further....

best regards, W.H.M.

I'm a Registered Histology Technician working in Research. I have a PI who wants to process whole Mice pups. He cuts the stomach from head to tail and fixes for a week, but when I processed them on my "Animal Organ" program on the VIP5, they were not processed properly. I'm thinking I need a longer run and wondered if anyone had experience processing whole Mice pups.

Dear Cynthia, was fascinated about your rapid answer here... Thank you. My rapid guess were:

since stomach is 'quite (densely) musculous" with different fiber directions as well as interspersed connective tissue it is a really challenging task. Excuse me that at this end only a retired natural scientist speaks out - who performed rat whole brain [1 d PN to 120 days of age) processing - choosing then the area around 3rd ventricle for resin embedding (and did finally serial semithin sectioning by myself). You are right in saying you might have needed a longer run with all your processing steps in the Sakura-Tissue-Tek VIP5 -processor...BUT: also I'd mention that in the 1970ies until end of my studies in 1980 I've learned a lot in (also manual!) tissue processing from an also registered Histotechnician in earlier times too, that there might be an "overfixation" of a certain tissue (re: the type of fixative effectively used, proper / AND: ...improper... application of the fixative, as well as the period the whole or eventually half-sliced organ / tissue was impregnated / imbibed / immersed... not to forget temperature issues, previous 'surface drying' issues during first handling, ...), resulting in a really "hard" and henceforward almost impenetrable bunch of tissue.

...So it might be an issue to (re-) evaluate the regime / processing steps your PI has made previously to your receiving the specimens in your lab... (misprocessing might not be YOUR fault! - since - believe me - as a former 'bench-worker' and - finally - "finisher" of 'academic projects' I know what PI's, or academic students, doctors from other facilities, etc. are able to do "wrong" without announcement! (;....||). Therefore: Best wishes for a lucky troubleshooting, cordially (if not welcome: respectfully) yours, Wolfgang

I have done my research with Westar albino rat pups. The pups were anesthetized by giving sodium pentothal depending on its body weight. An incision is made in the abdomen and thoracic region . The cranial cavity is also opened by cutting the skin and soft vault in a plus pattern and fixed in Bouin’s fluid and also in 4 % paraformaldehyde in 0.1 M phosphate buffered saline for 24 hours.

The fixed tissue is left in running water for a minimum period of three hours

and the tissue is processed with graded alcohol (30%, 50%, 70% and 90%) for dehydration and then proceeded with changes of absolute alcohol ( I, II and III). This is followed by three successive changes in xylene for clearing and finally infiltrated and embedded in paraffin wax (melting point- 58°C). Sections were taken at 5µ thickness and collected in albumin coated slide. The mounted sections are left overnight to dry and ready for staining procedure.

Hope my procedure may given an idea !!!

Thanks and best wishes

Wolfgang,

Thank you so much for your very helpful reply to my problem!! I really appreciate the time you took to help me, which what you said, did help very much!! I had told the PI to bisect the pup then put into fixative, which would greatly help the fixation process. I also told him, if he had any more in the future, I would process them for a longer program.

Again, thank you for your time!!!

Take care,

Cynthia

Govindarajan,

Thank you for your reply! I will suggest to the PI to use Bouin's for the fixative. Your processor solutions are what I use on my Automatic Tissue Processor, VIP5, I was really wanting times for each of those solutions, but thank you for reply.

Take care,

Cynthia