Elisa sandwich with Luminex beads - (Nov/23/2010 )
Hi everybody,
I am trying to setup a ELISA experiment using Luminex beads. For this, I followed a standard protocol: coupling of the capture antibody to the beads, incubation of the coupled bead with my samples, incubation with a biotinylated secondary antibody and finally, incubation with PE-streptavidin. The problem is that I am getting MFI values of 150-500. The background is consistently low however(>15)and I do see a difference in signal between positive and negative samples (say, 50-100 against 300-500). The point is that I still consider these values very low when comparing with published results (MFI around 3000).
Does anyone have experience with this kind of assays? Are my results still significative although the MFI values are that low?
Thanks in advance!
Please correct me if I am wrong...but is the Luminex technology basically for analysis of multiple analytes in a single sample? If you are trying to detect a single analyte is this the correct approach?
Chelo on Tue Nov 23 18:18:44 2010 said:
Hi everybody,
I am trying to setup a ELISA experiment using Luminex beads. For this, I followed a standard protocol: coupling of the capture antibody to the beads, incubation of the coupled bead with my samples, incubation with a biotinylated secondary antibody and finally, incubation with PE-streptavidin. The problem is that I am getting MFI values of 150-500. The background is consistently low however(>15)and I do see a difference in signal between positive and negative samples (say, 50-100 against 300-500). The point is that I still consider these values very low when comparing with published results (MFI around 3000).
Does anyone have experience with this kind of assays? Are my results still significative although the MFI values are that low?
Thanks in advance!
MFI values of 150-500 will do. It depends antigen concentration in the sample, your sample dilution and your incubation times. I have assays both with low MFI (50-600) and with higher MFI (5000-10000) values.