Team:EPF-Lausanne/Protocol/TrypanBlue

From 2012.igem.org

(Difference between revisions)
 
(11 intermediate revisions not shown)
Line 1: Line 1:
<noinclude>{{:Team:EPF-Lausanne/Template/Header}}</noinclude>
<noinclude>{{:Team:EPF-Lausanne/Template/Header}}</noinclude>
-
{{:Team:EPF-Lausanne/Template/ProtocolHeader| Trypan Blue Method | {{{1|}}}}}
+
{{:Team:EPF-Lausanne/Template/ProtocolHeader|Trypan Blue Method | {{{1|}}}}}
-
{| {{table}}
+
<noinclude>{{:Team:EPF-Lausanne/Template/SetTitle|Trypan Blue}}</noinclude>
-
| align="center" style="background:#f0f0f0;"|''''''
+
 
-
| align="center" style="background:#f0f0f0;"|'''Sampling'''
+
=== Sampling ===
-
| align="center" style="background:#f0f0f0;"|''''''
+
{| class="wikitable" style="text-align: center; color: black;"
-
| align="center" style="background:#f0f0f0;"|''''''
+
|Cell Density (10^6/ml)
-
| align="center" style="background:#f0f0f0;"|''''''
+
|Dilution       
-
| align="center" style="background:#f0f0f0;"|''''''
+
|PBS              (µl)
 +
|Cells            (µl)
 +
|Trypan Blue (µl)
|-
|-
-
| Dilutions||Cell Density ( 10^6/ml)||Dilution||PBS (µL)||Cells (µL)||Trypan Blue(µL)
+
|1 - 2
 +
|4
 +
|100
 +
|50
 +
|50
|-
|-
-
| ||1-2||4||100||50||50
+
|2 - 4.5
 +
|8
 +
|125
 +
|25
 +
|50
|-
|-
-
| ||2- 4.5||8||125||25||50
+
|4.5 - 7
 +
|12
 +
|120
 +
|15
 +
|45
|-
|-
-
| ||4.5 - 7||12||120||15||45
+
|> 7
 +
|16
 +
|137.5
 +
|12.5
 +
|50
|-
|-
-
| ||>7||16||137.5||12.5||50
 
|}
|}
-
 
+
#Take x µL of PBS in a 96-well plate.
-
#Take xµL ofPBS in 96 well plate
+
#Add the required volume of cell culture. Mix once.
-
#Add required volume of cell culture. Mix once
+
#Bring the plate back to the microscope, add Trypan Blue to the PBS + cell mixture just before counting the sample.  
-
#Bring the plate back to microscope, add trypan blue to the PBS + Cell mixture just before counting the sample.  
+
-
      Trypan Blue is toxic to cells. If left too long with trypan blue, even healthy cells will die.  
+
Trypan Blue is toxic to cells. If left too long with it, even healthy cells will die.
'''Calculation of LCD :'''
'''Calculation of LCD :'''
-
LCD = Cell Count/ ( 100* 4) * Dilution
+
LCD = Cell Count/(100*4)*Dilution
'''Tips :'''
'''Tips :'''
-
# Mix cells before sampling
+
# Mix cells before sampling.
-
# Take cell sample from top of the liquid
+
# Take the cell sample from the top of the liquid.
-
# Mix trepan blue into the PBS + cel solution slowly and well before loading
+
# Mix Trypan Blue in the PBS + cell solution slowly before loading.

Latest revision as of 00:31, 18 September 2012

Protocol: Trypan Blue Method

Sampling

Cell Density (10^6/ml) Dilution PBS (µl) Cells (µl) Trypan Blue (µl)
1 - 2 4 100 50 50
2 - 4.5 8 125 25 50
4.5 - 7 12 120 15 45
> 7 16 137.5 12.5 50


  1. Take x µL of PBS in a 96-well plate.
  2. Add the required volume of cell culture. Mix once.
  3. Bring the plate back to the microscope, add Trypan Blue to the PBS + cell mixture just before counting the sample.

Trypan Blue is toxic to cells. If left too long with it, even healthy cells will die.


Calculation of LCD :

LCD = Cell Count/(100*4)*Dilution

Tips :

  1. Mix cells before sampling.
  2. Take the cell sample from the top of the liquid.
  3. Mix Trypan Blue in the PBS + cell solution slowly before loading.


Wiki Links

Important pages

Pages

Retrieved from "http://2012.igem.org/Team:EPF-Lausanne/Protocol/TrypanBlue"