1. No cells or fluorescence is observed on the slide, is it possible that the antigen has been detached during the washing step?
If the washing protocol is carried out correctly this should not be observed. Please, review the washing procedure avoiding actions that may damage/detach the antigen:
• The washing solution preparation and storage should be carried out following the instructions for use.
• Only the wash solution included in the Vircell kit should be used. The use of another manufacturer's wash solution should be avoided.
• The washing timings indicated in the instructions for use should be strictly followed.
• The direct incidence of a PBS/water jet in the well should be avoided.
• Finally, slide overlapping should also be avoided.
2. The Rickettsia / Bartonella slides look different to what I would expect from other commercial brands.
As part of our production process of Rickettsia and Bartonella, we completely remove all traces of the Vero cells where they are grown in order to increase the specificity of these products. In most cases, the patient's serum reacts with the Vero cell remnants creating fluorescence that can be interpreted as a positive reaction. By eliminating these remains, we minimize the possibility of false positives. This is actually an advantage of our products over those of the competition.
The procedure should be reviewed to make sure that the instructions for use have been rigorously followed:
• The washing solution preparation and storage should be carried out according to the above-mentioned indications for use.
• The washing protocol should be checked. It is important to follow the specified washing timings and apply gentle agitation to ensure that all wells are washed uniformly. The slides should be rinsed with distilled water after each wash as residual salts of the wash solution can cause background reading.
• The incubation timings should also be checked since a longer incubation than recommended can produce background.
• Please, check that the wells have not dried out during incubation, especially if the background affects only some of them. It is recommended to use a humid chamber during incubation to prevent the wells from drying out completely.
4. I obtain negative results at lower dilutions, and positive results at higher dilutions, for the same sample, what could be the reason?
This problem could indicate desiccation of the globulin. Please make sure that the incubations of the sample and globulin take place in a humid chamber. The washing step should also be checked in order to make sure that it has been carried out correctly. On the other hand, some samples may present prozone effect, however these cases are very rare and occur only in sera with very high titers.
6. Why does Vircell include wells of Chlamydia trachomatis y Chlamydophila psittaci in the Chlamydophila pneumoniae de IFA slide?
The evaluation of the immunological response to Chlamydia, using the MIF technique, must be carried out over the three Chlamydia species simultaneously. This is a way of controlling cross reactivity; the differential responses to the three Chlamydia species will render the final diagnosis. Therefore, the kit may be used to detect responses to any of the species. The positive control is positive for the three species.