2024 Bioimaging (Stephens)
2024 Bioimaging (Stephens) kdorfman Mon, 08/12/2024 - 22:37HeLas for teaching microscopy
MEFs for CURE
day | date | prep |
---|---|---|
Tu | 9/3 | 12 coverslips: HeLa (microtubules, actin, DAPI) |
Th | 9/5 | (Transmitted light and alignment) |
Tu | 9/10 | (Cameras, SNR, calibration) |
Th | 9/12 | (NA and resolution) beads |
Tu | 9/17 | (Fluorescence) |
Th | 9/19 | photobleaching (need new slides?) |
Tu | 9/24 | IF: need fixed coverslips, 10, 20 Ab |
Th | 9/26 | view IF from previous class, cell culture (MEF 39) |
Tu | 10/1 | Live cell imaging and cell culture |
Th | 10/3 | Plasmid prep Eb1 into MEF 39 and cell culture Kate transfects, puts into live dishes |
Tu | 10/8 | Transfection imaging |
Th | 10/10 | midterm |
Tu | 10/15 | NO CLASS (Monday schedule) |
Th | 10/17 | Begin CURE |
Tu | 10/22 | |
Th | 10/24 | |
Tu | 10/29 | |
Th | 10/31 | |
Tu | 11/5 | NO CLASS (Election Day) |
Th | 11/7 | |
Tu | 11/12 | |
Th | 11/14 | |
Tu | 11/19 | |
Th | 11/21 | |
Tu | 11/26 | CURE written report due |
Th | 12/3 | (Michael) Neurobiology |
Tu | 12/5 | (Michael) Neurobiology |
Tu | 12/10 | Flexible last day of class |
HeLa for 1st day
HeLa for 1st day kdorfman Wed, 08/28/2024 - 20:33For "Learn your Microscope" 9/3
9 HeLa (Vikash) coverslips
Fixed in paraglut
Stained for alpha tubulin:
Stained for green actin
Mounted in DAPI
Note: Red tubulin was gorgeous! Let's see if it lasts longer than the FITC.
IF for 9/24
IF for 9/24 kdorfman Wed, 08/28/2024 - 20:47Fixed coverslips (MEF 39 from Drew's lab)
Fix in paraformaldehyde
primary | secondary |
---|---|
Lamin B1 (rabbit, 1:1,000) | green |
NPC (mouse, 1:2,000) | red |
Emerin (mouse, 1:1,000) (from Drew's lab) | red |
To make the double primaries:
Make 2x Lamin B1 (add 1 mL instead of 2 mL)
Make 2x NPC (follow label directions = 2x (should say 1:20)
Make 2x Emerin: 1:500
Mix 250 µL each
To make the double secondary:
- Make 2x goat anti rabbit green
- goat anti rabbit green aliquots need 1194 µL to be 1X
- make 2X by adding 597 µL PBS-Tw-Az BSA
- Make 2X goat anti mouse red:
- aliquot says add 500 µL to make 1X
- add 250 µL to make 2X (use 2 aliquots)
- Make 1x green + red by mixing
- 500 µL goat anti rabbit green with
- 500 µL goat anti mouse red
DAPI
Slides
Beakers
forceps
Humid chambers