Recombinant Mouse Anti-B. asper BaP1 Antibody (scFvBaP1) (CAT#: FAMAB-0027CQ)

Figure 1 scFvBaP1 recognition of BaP1 toxin or whole venom by ELISA.

Samples of purified BaP1 or crude B. asper venom were coated onto 96 well plates and incubated with scFvBaP1, MABaP1 or SUMO. After washing and incubation with anti-HIS antibody, the antigen-antibody complex formation was detected using anti-mouse IgG-HRP and revealed under suitable enzymatic condition.

Gomes, M., Alvarez, M. A., Quellis, L. R., Becher, M. L., de Andrade Castro, J. M., Gameiro, J., ... & de Oliveira Santos, M. (2019). Expression of an scFv antibody fragment in Nicotiana benthamiana and in vitro assessment of its neutralizing potential against the snake venom metalloproteinase BaP1 from Bothrops asper. Toxicon, 160, 38-46.

Figure 2 scFvBaP1 neutralization of the fibrinolytic activity induced by BaP1.

A sample of BaP1 toxin (5 μg) was incubated (30 min/37 centigrade) with MABaP1 (at 2.5:1 or 5:1, antibody: toxin molar ratio) or scFvBaP1 (20:1 or 10:1 M ratio). Samples were applied to agarose plates containing human fibrinogen clotted by bovine thrombin, incubated (37 centigrade, overnight), subsequently, the hydrolyzed area was measured and expressed in mm². *p < 0.05 compared to PBS group values, #p < 0.05 compared to BaP1.

Gomes, M., Alvarez, M. A., Quellis, L. R., Becher, M. L., de Andrade Castro, J. M., Gameiro, J., ... & de Oliveira Santos, M. (2019). Expression of an scFv antibody fragment in Nicotiana benthamiana and in vitro assessment of its neutralizing potential against the snake venom metalloproteinase BaP1 from Bothrops asper. Toxicon, 160, 38-46.

Figure 3 Ability of MABaP1 and scFvBaP1 to neutralize BaP1-induced hemorrhage.

Hemorrhage was induced by BaP1 (35 μg) intradermally injected into the dorsal skin of Swiss mice. To neutralize BaP1-induced hemorrhage, MABaP1 or scFvBaP1 was incubated with BaP1 (1.5:1 or 10:1 M ratio,respectively). Mice were killed 3 h after injection, the skin was removed, the area of the hemorrhagic spots was assessed and expressed in mm². *p < 0.05 compared to saline values, #p < 0.05 compared to BaP1.

Gomes, M., Alvarez, M. A., Quellis, L. R., Becher, M. L., de Andrade Castro, J. M., Gameiro, J., ... & de Oliveira Santos, M. (2019). Expression of an scFv antibody fragment in Nicotiana benthamiana and in vitro assessment of its neutralizing potential against the snake venom metalloproteinase BaP1 from Bothrops asper. Toxicon, 160, 38-46.

Figure 4 Ability of scFvBaP1 to neutralize BaP1-induced myonecrosis.

To evaluate the myotoxic activity and the neutralizing potential of scFvBaP1 on this activity, mice were injected (i.m.) with either 20 μg BaP1 alone or previously incubated with scFvBaP1 (10:1 M ratio). After 4 h, the animals were bled and the sera were assayed for creatine kinase activity. *p < 0.05 compared to PBS values, #p < 0.05 compared to BaP1.

Gomes, M., Alvarez, M. A., Quellis, L. R., Becher, M. L., de Andrade Castro, J. M., Gameiro, J., ... & de Oliveira Santos, M. (2019). Expression of an scFv antibody fragment in Nicotiana benthamiana and in vitro assessment of its neutralizing potential against the snake venom metalloproteinase BaP1 from Bothrops asper. Toxicon, 160, 38-46.

Figure 5 Ability of scFvBaP1 to neutralize BaP1-induced inflammatory cell migration and H₂O₂ production in the peritoneal cavity of mice.

Total peritoneal exudate cell (PEC) (A) and production of H₂O₂ by exudate cells (B) were evaluated. BALB/c mice were injected (i.p.) with either PBS, BaP1 (5 μg) or BaP1 previously incubated with scFvBaP1 (10:1 M ratio). Mice were killed 6 h after treatment and total cells were counted in a Neubauer chamber. For detection of H₂O₂, 2.5 × 10⁶ cells/mL were incubated in the presence or absence of PMA (10 ng/well) followed by horseradish peroxidase (HRPO)-mediated oxidation of phenol red, consequently, resulting in increased absorbance at 620 nm.

Gomes, M., Alvarez, M. A., Quellis, L. R., Becher, M. L., de Andrade Castro, J. M., Gameiro, J., ... & de Oliveira Santos, M. (2019). Expression of an scFv antibody fragment in Nicotiana benthamiana and in vitro assessment of its neutralizing potential against the snake venom metalloproteinase BaP1 from Bothrops asper. Toxicon, 160, 38-46.

Figure 6 Ability of scFvBaP1 to recognize and neutralize the biological activities of BnP1 (a P-I SVMP isolated from Bothrops neuwiedi venom).

To evaluate the scFvBaP1 recognition of BnP1, samples of this SVMP were used to coat plates that were incubated with scFvBaP1. After addition of anti-HIS antibody, antigeneantibody complex formation was detected using anti-mouse IgG-HRP followed by incubation with suitable substrate (A). To evaluate the ability of scFvBaP1 to neutralize fibrinolysis (B) induced by BnP1, 5 μg of BnP1 was incubated (30 min/37 centigrade) with scFvBaP1 (20:1 or 10:1 M ratio). Samples were applied to agarose plates containing human fibrinogen clotted by bovine thrombin. After incubation (37 centigrade, overnight), the hydrolyzed area was measured and expressed in mm². The inhibition of BnP1-proinflammatory activities by scFvBaP1 was evaluated in mice.

Gomes, M., Alvarez, M. A., Quellis, L. R., Becher, M. L., de Andrade Castro, J. M., Gameiro, J., ... & de Oliveira Santos, M. (2019). Expression of an scFv antibody fragment in Nicotiana benthamiana and in vitro assessment of its neutralizing potential against the snake venom metalloproteinase BaP1 from Bothrops asper. Toxicon, 160, 38-46.