Comparative analysis of cell immunohistochemistry results obtained using fresh or frozen cells. A and C, correspond to experiments done on fresh cells; and B and D, on frozen cells. In A and B, the fluorescence signal from HEK cells expressing AQP4-EGFP is shown in green (left column), and that from the secondary antibody bound to the NMO-IgG in red (middle column), while the merged image of both fluorescent signals is shown in yellow (right column). Large magnification images are boxed. Two patients’ serum were used in the analysis; C+, in which NMO-IgG antibodies had been confirmed previously, and C-, in which the antibodies were absent. C and D summarize the quantitative analysis. A significant difference (** p ≤ 0.01) with respect to C + serum was obtained as indicated. Error bars are ± SEM (n = 5).