McTips

McTips 2018 PDF (as of 201810002) go to: McTips 2018 download.

My McTips 2017 is available online at   https://works.bepress.com/gmcnamara/81/

McTips 2017 Direct download is    https://works.bepress.com/gmcnamara/81/download 

McTips 2018 PDF (as of 201810002) go to: McTips 2018 download.

https://works.bepress.com/gmcnamara/83

The McTips 2018 includes some of my thoughts with respect to Fast Photon Counting (FPC) to make fluorescence confocal microscopy both faster and more quantitative than is now practiced by most biomedical researchers (i.e. twiddle the HV gain and offset values until someone proves their boss' hypothesis ... especially when they are using 'Santa Crap' antibodies and don't bother with controls).

ThermoFisher Prolong Glass  (without DAPI)     is now the best choice, if imaging fixed specimens with oil immersion objective lens. 

Prolong Glass info states needs to cure for 30+ hours.

My advice:

* grow cells in imaging dishes (mattek or WPI-Inc ... or ibidi imaging quality coverglass chambers)

** at no time should cells be allowed to "air dry" = keep submerged.

* fix (i.e. formaldehyde), permeabilize if needed.

* immunofluorescence (i.e. http://www.nano-tag.com 2ndary nanobodies with each mouse mAb) ... can include DAPI and/or other counterstains here (example: fluorescent phalloidin).

* wash extensively (but quickly).

* "drip on" some Prolong Glass with imaging dish tilted, so that it forces aqueous media away ... pipet out the "run off", drop more (but not too much $) Prolong Glass ... goal is ~100% Prolong Glass, ~0% aqueous.

* allow to "cure" 30+ hours, in the dark, at room temperature, no lid, large volume of air (i.e. not small sealed box) to let volatiles escape.​ ... Probably simplest to go closer to 48 hours (and would be nice to be consistent in experiments).

 

20180803Fri ... connecting to our file server from Windows (win 7).

* ask George for the name of our file server - and please do not give out the name or IP address of our server.

* you are welcome to set up your own 'share drive to transfer your files (and can we please have 42 Terabytes of space on yours?).

Some Windows PC's are able to see our file server. Some are not. Today we -- "we" being 99% Jim Potter and 1% GM -- were able to trouble shoot the network acess issue. 

0. assumptions:

      (i) windows PC (win7 or ideally win10)

      (ii) plugged into the JHU SOM network (Ethernet cable).

      (iii) you have administrator privileges on the PC (does not need to be 'Administrator' login name).

1. Use JHARS to connect to JHU network (if you have not already done so) https://jhars.nts.jhu.edu/

  1a. after setting  up (or confirming) JHARS, probably useful to power off the PC, wait a few seconds, then power up and log in.

2. enable all the items in the Local Area Connection Properties dialog box (it is probably ok to enable more, but at minimum you need IPv6 and IPv4 and probably more).

3. using CMD prompt -> IPconfig / all (2nd screen shot below) ... see that DHCP Server 10p15.76.226

local network

 

Windows Start menu ... cmd ... ipconfig / all

    ==> DHCP Server 10.15.176.226

    ==> Subnet mask 255.255.255.0     (if this is not correct, you may not be able to see JHU network at all!).

ipconfig all

 

 

 

 

 
 
 
 
 
 
 

 

 

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