Difference between revisions of "Altman:Protocols/Single Molecule Assays/gliding filament"

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==Making Nitrocellulose Coverslips==
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==Gliding Filament Assay==
  
 
''Typical flow-through volumes are 10-15 μL''
 
''Typical flow-through volumes are 10-15 μL''
  
 
1. Make a flow cell with a nitrocellulose-coated coverslip
 
1. Make a flow cell with a nitrocellulose-coated coverslip
 +
 
2. Flow in α-GFP/10
 
2. Flow in α-GFP/10
 +
 
3. Incubate 2 minutes
 
3. Incubate 2 minutes
 +
 
4. Flow in ABSA
 
4. Flow in ABSA
 +
 
5. Incubate 2 minutes
 
5. Incubate 2 minutes
 +
 
6. Flow in motor dilution
 
6. Flow in motor dilution
 +
 
7. Incubate 2 minutes
 
7. Incubate 2 minutes
 +
 
8. Flow in ABSA
 
8. Flow in ABSA
 +
 
9. Flow in Actin dilution  
 
9. Flow in Actin dilution  
 +
 
10. Incubate 2 minutes
 
10. Incubate 2 minutes
 +
 
11. Flow in GO-Juice
 
11. Flow in GO-Juice
  

Revision as of 12:50, 15 June 2013

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Gliding Filament Assay

Typical flow-through volumes are 10-15 μL

1. Make a flow cell with a nitrocellulose-coated coverslip

2. Flow in α-GFP/10

3. Incubate 2 minutes

4. Flow in ABSA

5. Incubate 2 minutes

6. Flow in motor dilution

7. Incubate 2 minutes

8. Flow in ABSA

9. Flow in Actin dilution

10. Incubate 2 minutes

11. Flow in GO-Juice


Recipes

50 uL Buffer #1
50x GOC stock 2 μL
100x CPK stock 1 μL
100x ATP stock 1 μL
AB 46 μL


50 μL Buffer #2
50x Glu/B stock 2 μL
100x PCr stock 1 μL
AB 47 μL


20 μL GO Juice
Buffer #1 10 μL
Buffer #2 10 μL