Recombinant Mouse Anti-C. neoformans GXM Antibody (3E5) (CAT#: FAMAB-0042CQ)

Figure 1 Changes in hematocrit after administration of MAbs of various isotypes.

Infected Swiss-Webster mice were given 0.5 mg intraperitoneally of one of following MAbs: 3E5 IgG1, 3E5 IgG3, 2D10 IgM, or 18G9 IgA. Symbols represent mean hematocrit levels. Brackets denote SD. n = no. of mice in each group. Hematocrit before MAb administration was measured in 46 mice. Effect of intervention is compared with mean hematocrit of these 46 mice

Lendvai, N., & Casadevall, A. (1999). Monoclonal Antibody—Mediated Toxicity in Cryptococcus neoformans Infection: Mechanism and Relationship to Antibody Isotype. The Journal of infectious diseases, 180(3), 791-801.

Figure 2 Histopathology of mice given MAbs on day 10 of infection.

A, C, E, G, tissue from infected mouse given MAb 3E5 IgG1. B, D, F, H, tissue from infected mouse given MAb 3E5 IgG3. Original magnifications: A, B, kidney (×10); C, D, kidney (×20); E, F, liver (×10); G, H, lung (×10). Red blood cells seen distending vessels and glomeruli of kidney in mice treated with 3E5 IgG1 are largely absent in mice given MAb 3E5 IgG3 with no clinical signs of toxicity. Hematoxylin-eosin stain.

Lendvai, N., & Casadevall, A. (1999). Monoclonal Antibody—Mediated Toxicity in Cryptococcus neoformans Infection: Mechanism and Relationship to Antibody Isotype. The Journal of infectious diseases, 180(3), 791-801.

Figure 3 Immunohistochemical localization of glucuronoxylomannan (GXM) in livers of mice given MAbs on day 10 of infection.

Infected mouse given MAb 3E5 IgG1 (A, C) and infected mouse given MAb 3E5 IgG3 (B, D). Original magnifications: ×20 (A, B); ×100 (C, D). GXM appears to be similarly localized, in part intracellularly in both mice, despite treatment of one with "toxic" and other with "nontoxic" isotype.

Lendvai, N., & Casadevall, A. (1999). Monoclonal Antibody—Mediated Toxicity in Cryptococcus neoformans Infection: Mechanism and Relationship to Antibody Isotype. The Journal of infectious diseases, 180(3), 791-801.

Figure 4 ELISA

A, binding of changing concentrations of mAbs 3E5 IgA and 3E5 IgE to immobilized GXM. Isotype-specific secondary Abs were used as well as anti-κ secondary Ab. The inset represents the schematic for the ELISA configuration. B, competition ELISA of mAb 3E5 IgE with changing concentrations of mAb 3E5 IgA to immobilized GXM. Secondary Ab used is goat anti-mouse-alkaline phosphatase (GAM-AP) IgE. The schematic represents the ELISA configuration.

Janda, A., Eryilmaz, E., Nakouzi, A., Pohl, M. A., Bowen, A., & Casadevall, A. (2015). Variable region identical IgA and IgE to Cryptococcus neoformans capsular polysaccharide manifest specificity differences. Journal of Biological Chemistry, 290(19), 12090-12100.

Figure 5 Immunofluorescence of strain 24067 stained with mAbs 3E5 IgA and 3E5 IgE.

Janda, A., Eryilmaz, E., Nakouzi, A., Pohl, M. A., Bowen, A., & Casadevall, A. (2015). Variable region identical IgA and IgE to Cryptococcus neoformans capsular polysaccharide manifest specificity differences. Journal of Biological Chemistry, 290(19), 12090-12100.

Figure 6 A, J774 cells and C. neoformans were used to study the phagocytosis efficacy of the mAbs. B, survival of Balb/c mice with either NSO or the two variants of 3E5 mAb. The Abs were administered 2 h before infection.

Janda, A., Eryilmaz, E., Nakouzi, A., Pohl, M. A., Bowen, A., & Casadevall, A. (2015). Variable region identical IgA and IgE to Cryptococcus neoformans capsular polysaccharide manifest specificity differences. Journal of Biological Chemistry, 290(19), 12090-12100.

Figure 7 ELISA

(A and B) ELISAs of 3E5 IgG (IgG1, IgG2a, IgG2b, and IgG3) and of 4H3 IgG (IgG1, IgG2b, and IgG3) MAb binding to GXM antigen, respectively; (C and D) competition assays. The details of each ELISA are described in Results and are diagrammed in the insert of each panel.

Yuan, R. R., Spira, G., Oh, J., Paizi, M., Casadevall, A., & Scharff, M. D. (1998). Isotype switching increases efficacy of antibody protection against Cryptococcus neoformans infection in mice. Infection and immunity, 66(3), 1057-1062.

Figure 8 Survival of A/JCr mice infected with C. neoformans after the admin istration of IgG3 isotype switch variant MAbs.

A dose of 1.0 mg of each antibody was given i.p. 24 h prior to i.v. challenge with 2 × 10⁶ C. neoformans cells. (A)Average survival rates ± standard deviations for the 3E5 IgG3, IgG1, IgG2b, and IgG2a and PBS groups were 11.6 ± 0.5, 14.4 ± 2.6, 14.0 ± 1.7, 15.8 ± 2.6, and 12.1 ± 1.0 days, respectively. IgG1, IgG2b, and IgG2a MAbs prolonged animal survival significantly (P < 0.003), whereas the IgG3 MAbs showed a trend toward decreased survival that was not statistically significant (P > 0.18). (B) Average survival rates ± standard deviations for the 4H3 IgG3, IgG1, and IgG2b and PBS groups were 5.8 ± 3.8, 16.3 ± 3.2, 17.9 ± 2.8, and 14.1 ± 3.1 days, respectively. Administration of 4H3 IgG3 MAb reduced survival significantly (P < 0.0001). However, the 4H3 IgG1 switch variant showed no effects on survival (P > 0.14), and 4H3 IgG2b prolonged animal survival (P < 0.01).

Yuan, R. R., Spira, G., Oh, J., Paizi, M., Casadevall, A., & Scharff, M. D. (1998). Isotype switching increases efficacy of antibody protection against Cryptococcus neoformans infection in mice. Infection and immunity, 66(3), 1057-1062.

Figure 9 Survival of A/JCr mice infected with C. neoformans after two administrations of the 3E5 IgG3 MAb.

A dose of 1 mg of the 3E5 MAb was given i.p. 24 h prior to and 3 days after i.v. challenge with 2 × 10⁶ C. neoformans cells. Average survival rates ± standard deviations for the groups given IgG3 once and twice and the PBS group were 11.5 ± 1.1, 9.0 ± 3.3, and 14.1 ± 1.2 days, respectively. Administration of two doses of IgG3 3E5 reduced survival relative to one dose (P < 0.002).

Yuan, R. R., Spira, G., Oh, J., Paizi, M., Casadevall, A., & Scharff, M. D. (1998). Isotype switching increases efficacy of antibody protection against Cryptococcus neoformans infection in mice. Infection and immunity, 66(3), 1057-1062.

Figure 10 Effects of 3E5 and 4H3 MAb switch variants on phagocytosis by A/JCr bronchoalveolar macrophages in the presence or absence of IFN-γ.

All MAbs promoted phagocytosis, and phagocytosis was significantly greater in cells treated with IFN-γ (P < 0.01).

Yuan, R. R., Spira, G., Oh, J., Paizi, M., Casadevall, A., & Scharff, M. D. (1998). Isotype switching increases efficacy of antibody protection against Cryptococcus neoformans infection in mice. Infection and immunity, 66(3), 1057-1062.