Article Figures & Data
Figures
- FIG. 1.
Expression and purification of rTbps. (A) Coomassie blue-stained SDS-polyacrylamide gel containing rTbpA protein. Lane 1 contains the soluble fraction after detergent solubilization of induced E. coli. Lanes 2 through 7 represent rTbpA purification fractions. Lane 2 contains the column flowthrough fraction after overnight incubation with a human transferrin-bound affinity column. Lane 3 contains the wash fraction. Lanes 4 through 7 contain purified rTbpA column elution fractions. Molecular weight standards (MWM) are indicated at the left. (B) Western blot of the above SDS-PAGE probed with an anti-TbpA antibody. The positions of molecular weight standards are indicated on the left. (C) Solid-phase transferrin binding assay of purified rTbpA probed with HRP-transferrin (1 μg/ml). Lane 1 contains duplicate spots of purified rTbpA. Lane 2 contains duplicate spots of buffer only. (D) Coomassie blue-stained SDS-PAGE containing rTbpB protein. Lane 1 contains a whole-cell lysate of IPTG-induced E. coli. Lane 2 contains the soluble fraction of IPTG-induced, detergent-solubilized E. coli. Lanes 3 through 6 represent rTbpB purification fractions. Lane 3 contains the flowthrough after overnight incubation with a nickel-affinity resin. Lane 4 contains the wash fraction. Lanes 5 through 6 contain purified rTbpB fractions. Molecular weight standards (MWM) are indicated at the left. (E) Western blots of purified rTbpB. Panel 1 was probed with an anti-TbpB antibody. Panel 2 was probed with HRP-transferrin (1 μg/ml). (F) Solid-phase transferrin binding assay of purified rTbpB probed with HRP-transferrin (1 μg/ml). Lane 1 contains duplicate spots of rTbpB. Lane 2 contains duplicate spots of buffer only.
- FIG. 2.
IgG, IgM, and IgA antibody responses in serum. (A) Antibody responses against rTbpA. (B) Antibody responses against rTbpB. The infected groups are divided into those with a first gonococcal infection (filled symbols) and those with at least one prior episode of gonorrhea (open symbols). The control group is divided into volunteer lab personnel (filled symbols) and individuals attending an STD clinic who were culture negative with no previous history of gonococcal disease (open symbols). The horizontal bars indicate median values. Differences between the groups were compared with the Mann-Whitney U test or the Kolmogorov-Smirnov test where appropriate. Note the logarithmic scales.
- FIG. 3.
Western blot analysis of patient sera. (A) Western blot of a total-membrane preparation of iron-stressed N. gonorrhoeae probed with patient sera diluted 1:150. (B) Western blot of a total membrane preparation of iron-stressed N. meningitidis probed with the same dilution of patient sera as in panel A. (C) Western blot of purified gonococcal TbpA and TbpB probed with patient sera diluted at 1:100. Lanes 1 and 16 are negative controls with no antibody. Lane 2 was probed with an anti-TbpA antibody. Lanes 3 through 5 are female control sera. Lanes 6 and 7 are male control sera. Lanes 8 and 9 are sera from females infected with gonorrhea for the first time. Lanes 10 and 11 contain sera from females with at least one prior gonococcal infection. Lane 12 contains sera from a male infected with gonorrhea for the first time. Lanes 13 and 14 contain sera from males with at least one prior gonococcal infection. Lane 15 was probed for TbpB by using HRP-transferrin.
- FIG. 4.
Comparison of serum IgG responses to rTbpA, rTbpB, and tetanus toxoid. Patients' identification numbers and genders are indicated on the x axis. Antibodies to rTbpA or rTbpB were not measured in the control serum (C+/M). Note the logarithmic scale.
- FIG. 5.
Longitudinal serum antibody response. Serum antibody responses to rTbpA and rTbpB were detected from an infected male individual following four separate episodes of gonorrhea over a 6-month period. Following the initial infection, subsequent infections occurred at 1, 3, and 6 months. Note the logarithmic scale.
Tables
- TABLE 1.
Total genital tract immunoglobulin levels from uninfected and infected volunteers
Sample type and antibody class Median Ig levels (μg/ml) (range [n]) No previous history Current Infection Cervical mucus IgG 15.6 (0.1-178.6 [14]) 6.1 (0.3-89.8 [11]) IgM 0.3 (0.0-64.0 [14]) 4.6 (0.1-34.5 [11]) IgA 9.6 (0.2-121.5 [14]) 8 (0.0-88.5 [11]) Seminal plasma IgG 75.7 (49.5-187.0 [4]) 75.5 (34.6-727.5 [9]) IgM 1.2 (0.4-7.5 [4]) 3.5 (1.1-47.0 [9]) IgA 79.3 (54.4-80.1 [4]) 42.5a (27.0-348.7 [9]) ↵a P < 0.05 compared with results for uninfected controls (as determined by the Kolmogorov-Smirnov test).
