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ELISAS: U-bottom plates vs. flat bottom? - (Nov/18/2005 )

Ok, so I spend two days running a 10 plate ELISA (Costar's U-bottom) and it didn't work.
I ran the same assay on flat bottom plates a couple of weeks ago and it worked out fine.
I know I didn't miss anything in my assay so I think I made the big mistake using U-bottom plates (at least I think, perhaps my Ab did not bind).

Anyway, does anyone know the advantages/disadvantages to using U-bottom vs. flat plates on ELISAS?

-sharpie-

Hi Sharpie
Were your U well plates designed for ELISA. Not many types are. Costar do a range of U well plates that have no pre-treatment at all eg for red cell assays. They probably do special binding plates for ELISA but check which ones you have. Secondly U well plates have 2 advantages in that an increased surface area at the bottom of the well for antibody/antigen binding and the dynamics of well washing are better. That said the difference between U wells and flat wells (ELISA specific) is not that great and I have developed plenty of assays with Flat bottomed plates. Most important in choosing plates is the hydrophobic/hydrophilic nature of the plates. Some companies such as Nunc, Griener and Dynex provide plates of low, medium and high binding capacities. You could get free samples and give them a try if you ever need to improve your assay.
Regs
parky


QUOTE (sharpie @ Nov 18 2005, 03:29 PM)
Ok, so I spend two days running a 10 plate ELISA (Costar's U-bottom) and it didn't work.
I ran the same assay on flat bottom plates a couple of weeks ago and it worked out fine.
I know I didn't miss anything in my assay so I think I made the big mistake using U-bottom plates (at least I think, perhaps my Ab did not bind).

Anyway, does anyone know the advantages/disadvantages to using U-bottom vs. flat plates on ELISAS?

-Parky-

QUOTE (Parky @ Nov 18 2005, 08:16 AM)
Hi Sharpie
Were your U well plates designed for ELISA. Not many types are. Costar do a range of U well plates that have no pre-treatment at all eg for red cell assays. They probably do special binding plates for ELISA but check which ones you have. Secondly U well plates have 2 advantages in that an increased surface area at the bottom of the well for antibody/antigen binding and the dynamics of well washing are better. That said the difference between U wells and flat wells (ELISA specific) is not that great and I have developed plenty of assays with Flat bottomed plates. Most important in choosing plates is the hydrophobic/hydrophilic nature of the plates. Some companies such as Nunc, Griener and Dynex provide plates of low, medium and high binding capacities. You could get free samples and give them a try if you ever need to improve your assay.
Regs
parky


Hey there, My guess is that they were not designed for ELISAs (well, y assay didn't work).

I was given a nice set of NUNC plates (flat-bottom) designed for ELISAs so I'll go ahead and coat those over the weekend.

so now I know! Der!

Thanks!

-sharpie-

Hi there,
our lab technician told me that besides some plates being specifically designed for ELISA coating, you should also take into account that round bottom plates have a different refraction compared to flat bottom plates. Whether this will influence the read out I do not know for sure, but I can imagine it is possible.

Kind Regards,
Walter

-Walter-