Fig 1.
Evaluation of different codes of microspheres (#012, #035 and #070).
A: Microspheres (#012, #035 and #070) coupled with 5 μg/mL of A1 (black bars) and A4 (dashed bars) conjugates. Fluorescence intensity represented by the means of three geometric means ± standard deviation. B: Representative histograms of one experiment. PE-fluorescence intensity (X axis) vs. count (Y axis). One way ANOVA was performed for group comparison and significant differences were observed among all groups (p≤0.05).
Fig 2.
Evaluation of the best conjugate concentration and dilution.
A: Anti-IgG-PE concentration test (0.5, 1, 2.5 and 5 μg/mL). * = statistically different value when compared to the lowest concentration (p≤0.05). A1- black bars, A4- dashed bars and B2- white bars. B: Anti-HBSAg-PE dilution test (1:3000, 1:1000 and 1:300). C- black bars and D- dashed bars. One way ANOVA was performed for group comparison * = statistically different value among concentrations (p≤0.05). Data are expressed as mean of three geometric means ± standard deviation.
Fig 3.
Staining profile of anti-IgG-PE conjugates.
A: Representative histograms of PE-fluorescence intensity (X axis) vs. count (Y axis). 0.5 μg/mL = white peak and 5 μg/mL = black peak. B: Coefficient of variation of 0.5 μg/mL (white bars) and 5 μg/mL (black bars). Data expressed as the means of CVs from of three tests ± standard deviation. T test was performed for group comparison. An asterisk indicates a significantly different value assumed at p≤0.05.
Fig 4.
QFCM evaluation of anti-IgG-PE and anti-HBSAg-PE conjugates.
A: Histogram of PE-fluorescence intensity (X axis) vs. count (Y axis) of MESF kit (populations blank, bead #1, bead #2, bead #3 and bead #4). B: Geometric means of MESF kit beads along the months. C: MESF values of A1, A2, A3, A4, B1 and C conjugates along the months. A4 conjugate was coupled later than the other conjugates and its intensities evaluation started at month 4.
Table 1.
FITC and PE detection after anti-IgG-FITC staining.
Fig 5.
Conjugate stability study during 18 months.
A: A1, A2, A3, A4 and MESF bead #4 fluorescence intensities during the months of analysis. B: B1, B2, C, D and MESF bead #4 fluorescence intensities during the months of analysis. C: Conjugates histograms of PE-fluorescence intensity (X axis) vs. count (Y axis) on the first month (black peak) and last month of analysis (white peak). MESF bead #4 was included as an example of the MESF kit used to control instrument performance during the 18 months of evaluation. The A4 conjugate was coupled later than the other conjugates and its intensities evaluation started at month 4.