CONFOCALMICROSCOPY Archives

January 1997

CONFOCALMICROSCOPY@LISTS.UMN.EDU

Options: Use Monospaced Font
Show Text Part by Default
Condense Mail Headers

Message: [<< First] [< Prev] [Next >] [Last >>]
Topic: [<< First] [< Prev] [Next >] [Last >>]
Author: [<< First] [< Prev] [Next >] [Last >>]

Print Reply
Sender:
Confocal Microscopy List <[log in to unmask]>
Subject:
From:
Date:
Tue, 21 Jan 1997 12:10:39 -0500
Comments:
Reply-To:
Confocal Microscopy List <[log in to unmask]>
Parts/Attachments:
text/plain (45 lines)
For fixing head lice at different stages of oviposition, I used liquid
"canned air" to stop them in their tracks, then tipped in an excess of hot
alcoholic Bouin (or Dubosq Brasil as we entomologists called it).  They were
kept at 60 degrees for 1 hour, then transferred to alcohol for dehydration
(Automatic embedder, 45 minute changes at 80%, 96% then 100% ethanol),
clearing in toluene (2 x 45 min) and embedding in soft (mp 60 degrees)
paraffin wax (2 x 45 min).  This worked better than formaldehyde,
gluteraldehyde, Susa, regular Bouin, or Carnoy for histological preservation.
 Fixing punctured insects or removed abdomens gave no benefit in
preservation.  I understand Picric acid fixatives preserve antigenicity at
least as well as formalin.

For dissecting out the reproductive system of head lice, I sprinkled them on
a wax dish and used a soldering iron to melt in front of their head so they
are trapped thorax deep in wax.  By slicing off the tip of the abdomen with a
mounted razor, all the innards can be squeezed out with fine angled forceps
and fixed in formaldehyde.  This got my dissection time down to a few seconds
each and allowed me to do SEM of the internal organs.

For making transparent whole mounts of fleas, the insects can be collected in
30% alcohol.  I can't lay my hands on the exact protocol, but know it came
from one of Mirriam Rothschild's old papers.  The short cut was to put one
pellet of KOH in an eppendorf with a ml of water.  When dissolved, I added
the fleas and kept it in a 60 degree incubator for I think 4 hours.  The
longer you leave them, the lighter they get.  Embedding was done through
alcohol, then oil of clove (try Xylene), then DPX.

Good luck

David Carter
oQQQQQ@

Meridian Instruments Inc.

(Purveyors of premium confocal laser
scanning microscopes and workstations)

2310 Science Parkway
Okemos Michigan
48864 USA

Tel. 517 349-7200
Fax 517 349-5967
E-mail [log in to unmask]

ATOM RSS1 RSS2