ant surface stainig

[Doreen MILIUS]

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Dear list members,

we are trying to perform an unspecific fluorescence surface staining of whole ants to see the structure of the cuticle. We have tried water-soluble protein binding dyes like NHS-ster Dylight 549 which didn't work so well and are now thinking about trying out hydrophobic dyes that stain membranes unspecifically. Does anyone have experience with staining insect surfaces and could give a suggestion. This would be of great help.

Thanks a lot, Doreen


- - -
Doreen Milius

Bioimaging Facility
Institute of Science and Technology Austria
Am Campus 1
A-3400 Maria Gugging

[mcammer]

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What about airbrushing a thin fluorescent paint onto them, kind of like rotary shadowing for EM?
-Michael C.

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Doreen MILIUS
Sent: Monday, July 16, 2012 10:29 AM
To: [hidden email]
Subject: ant surface stainig

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Dear list members,

we are trying to perform an unspecific fluorescence surface staining of whole ants to see the structure of the cuticle. We have tried water-soluble protein binding dyes like NHS-ster Dylight 549 which didn't work so well and are now thinking about trying out hydrophobic dyes that stain membranes unspecifically. Does anyone have experience with staining insect surfaces and could give a suggestion. This would be of great help.

Thanks a lot, Doreen


- - -
Doreen Milius

Bioimaging Facility
Institute of Science and Technology Austria Am Campus 1
A-3400 Maria Gugging

[Oshel, Philip Eugene]

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Doreen,

A couple of questions: from your email, I infer that you want to
image the exterior surface of the intact ant. So:
The ants' cuticles will be covered by a waxy epicuticle, so a
oleophilic fluorescent stain would be needed, or a hexane wash to
remove the wax. Amount of wax is more-or-less dependent on ecology -
how xeric the ant's environment is, etc.

Insect cuticle is typically autofluorescent; have you tried imaging
for that? Or is it too dim for your needs?

Why not scanning electron microscopy? This is the simplest and best
way to get exterior morphology of arthropods generally and insects in
specific**. Are you trying to keep the ants alive? Environmental SEM
should allow this, as well as imaging without sputter-coating or the
like. This will image much finer details of the cuticle than you will
get using fluorescence microscopy.

Phil
**There is an excellent volume, "Identification Guide to the Ant
Genera of the World" by Barry Bolton that has SEM images of all the
genera, and good tips for mounting the ants for the SEM.

>*****
>To join, leave or search the confocal microscopy listserv, go to:
>http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
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>
>Dear list members,
>
>we are trying to perform an unspecific fluorescence surface staining
>of whole ants to see the structure of the cuticle. We have tried
>water-soluble protein binding dyes like NHS-ster Dylight 549 which
>didn't work so well and are now thinking about trying out
>hydrophobic dyes that stain membranes unspecifically. Does anyone
>have experience with staining insect surfaces and could give a
>suggestion. This would be of great help.
>
>Thanks a lot, Doreen
>
>
>- - -
>Doreen Milius
>
>Bioimaging Facility
>Institute of Science and Technology Austria
>Am Campus 1
>A-3400 Maria Gugging

--
Philip Oshel
Microscopy Facility Supervisor
Biology Department
024C Brooks Hall
Central Michigan University
Mt. Pleasant, MI 48859
[phone]

[Kilgore, Jason-2]

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I'm surprised the NHS ester didn't work well, since it should label all proteins.  In what way did it not work?  Ununiform labeling?  Too dim?

Jason

Jason A. Kilgore
Technical Application Scientist
Molecular Probes Labeling and Detection Technologies
Cells Systems Division
 
T 1 800 955 6288 then option 4, then option 6,  or  541 335 0353 . F 541 335 0238
29851 Willow Creek Rd . Eugene . OR . 97402-9132 . United States
www.invitrogen.com/technicalsupport

 




-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Doreen MILIUS
Sent: Monday, July 16, 2012 7:29 AM
To: [hidden email]
Subject: ant surface stainig

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Dear list members,

we are trying to perform an unspecific fluorescence surface staining of whole ants to see the structure of the cuticle. We have tried water-soluble protein binding dyes like NHS-ster Dylight 549 which didn't work so well and are now thinking about trying out hydrophobic dyes that stain membranes unspecifically. Does anyone have experience with staining insect surfaces and could give a suggestion. This would be of great help.

Thanks a lot, Doreen


- - -
Doreen Milius

Bioimaging Facility
Institute of Science and Technology Austria
Am Campus 1
A-3400 Maria Gugging

[Cameron Nowell]

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Hi Doreen,

I have gotten some great images of insects with my son playing around in
the facility during school holidays:)

If you have a confocal and can configure it for reflective imaging (i.e
detection of the reflected laser light coming back off the surface) you
will get some very nice and detailed images without the need for any dye
or image preparation (I just embedded the bugs in some low melt agarose
to keep them still).  Just be careful not to use too much laser power as
you could damage your detectors.


Cheers

Cam


Cameron J. Nowell
Microscopy Manager
Centre for Advanced Microscopy
Ludwig Institute for Cancer Research Melbourne - Parkville Branch
PO Box 2008
Royal Melbourne Hospital
Victoria, 3050
AUSTRALIA
Office: [phone]
Mobile: [phone]
Fax: [phone]
Facility Website
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-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]]
On Behalf Of Doreen MILIUS
Sent: Tuesday, 17 July 2012 12:29 AM
To: [hidden email]
Subject: ant surface stainig

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Dear list members,

we are trying to perform an unspecific fluorescence surface staining of
whole ants to see the structure of the cuticle. We have tried
water-soluble protein binding dyes like NHS-ster Dylight 549 which
didn't work so well and are now thinking about trying out hydrophobic
dyes that stain membranes unspecifically. Does anyone have experience
with staining insect surfaces and could give a suggestion. This would be
of great help.

Thanks a lot, Doreen


- - -
Doreen Milius

Bioimaging Facility
Institute of Science and Technology Austria Am Campus 1
A-3400 Maria Gugging