Materials and Reagents

1. Caco2 cell line (ATCC, catalog number: HTB-37 )
   
2. MEM (Mediatech, Cellgro^®, catalog number: 10-010-CV , or equivalent)
   
3. Glutamax (Life Technologies, Gibco^®, catalog number: 35050-061 , or equivalent),
   
4. Sodium pyruvate (Life Technologies, Gibco^®, catalog number: 11360-070 , or equivalent)
   
5. 10% heat-inactivated FBS
   
6. Porcine type 1x trypsin (Sigma-Aldrich, catalog number: T-0303 , or equivalent)
   
7. MgCl₂
   
8. Sucrose
   
9. Phosphate buffered saline (PBS)
   
10. Bovine serum albumin Fraction V (BSA) (Life Technologies, Gibco^®, catalog number: 15260-037 )
    
11. Formaldehyde (Polysciences, catalog number: 18814 )
    
12. TritonX-100 (Sigma-Aldrich, catalog number: 9002-93-1 )
    
13. Normal goat serum (Sigma-Aldrich, catalog number: 191356 )
    
14. 4',6-diamidino-2-phenylindole (DAPI) (Life Technologies, Molecular Probes^®, catalog number: D1306 )
    
15. Astrovirus 8E7 mouse monoclonal antibody (hybridoma cell line) (ATCC, catalog number: HB-11945 , or equivalent)
    
16. Fluorescent-conjugated anti-mouse IgG antibody [Life Technologies, Alexa Fluor^® 488 Goat Anti-Mouse (H+L) Antibody, catalog number: A11001 , or equivalent]
    
17. Astrovirus (not commercially available)
    
18. Bleach (clorox or stored brand equivalent)
    
19. Virkon S (DuPont)
    
20. Caco2 cell culture medium (see Recipes)
    
21. Serum-free (SF) Caco2 medium (see Recipes)
    
22. TN buffer (see Recipes)
    

Equipment

1. Biosafety cabinet
   
2. Gloves
   
3. Labcoat
   
4. Beckman ultracentrifuge tubes (ultra-clear 9/16 x 3 ½ in) (Beckman Coulter, catalog number: 344059 )
   
5. Beckman ultracentrifuge (that can reach 34,000 rpm and 4 ºC)
   
6. SW41 rotor
   
7. Pierce Extra-Strength Slide-A-Lyzer 10K molecular weight cassette (Pierce, catalog number: 66383 or 66380 )
   
8. Laminar flow hood
   
9. Syringe
   
10. Liquid nitrogen
    
11. Water bath
    

Procedure