Pnpp one component substrate

Manufactured by Southern Biotech

PNPP one component substrate is a laboratory reagent used as a substrate for various enzymatic assays. It is a colorless solution that undergoes a color change reaction when catalyzed by enzymes, allowing for the quantification of enzyme activity.

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Lab products found in correlation

Bsa fraction 5, by Merck Group (3 mentions) Imject alum, by Thermo Fisher Scientific (3 mentions) 96 well elisa plate, by Thermo Fisher Scientific (2 mentions) Np40 ficoll, by Biosearch Technologies (1 mentions)

3 protocols using pnpp one component substrate

Immunogenicity of Recombinant gp120 from Different Cell Lines

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Each group of C57Bl/6 mice was immunized with gp120 prepared from either 293F or
CHO-S cells. The recombinant gp120 (50 µg) was mixed with Imject Alum (Thermo
Fisher Scientific) and injected subcutaneously. Mice were boosted with same dose of
antigen at the indicated days along with Alum. Serum gp120 specific Ig levels in
these mice were analyzed by ELISA. Briefly, 96 well ELISA plates (Nalgene, Nunc) were
coated with gp120 (0.8 μg/well) overnight at 4°C, washed and blocked
with 1% BSA fraction V (Sigma–Aldrich), serum titers were then added to the
plates and incubated 4 hr at 4°C. After washing alkaline phosphatase-labeled
goat anti-mouse IgM or IgG isotype specific antibodies were added for 2 hr at room
temperature (SouthernBiotech, Birmingham, AL). After washing, PNPP one component
substrate (SouthernBiotech) was used to detect the amount of antibody bound.

Park C., Arthos J., Cicala C, & Kehrl J.H. (2015). The HIV-1 envelope protein gp120 is captured and displayed for B cell recognition by SIGN-R1+ lymph node macrophages. eLife, 4, e06467.

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Immunization and Antibody Detection

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Wild-type and mutant 6- to 8-wk-old mice were immunized with either sheep RBCs, NP₃₅-KLH, or NP₄₀-Ficoll. For the sheep RBC immunizations 200 μl of 10% solution of sheep RBCs (Lonza Walkerville, Inc.) was given by intraperitoneal injection. NP₃₅-KLH (Biosearch Technology) was mixed with Imject® Alum (Thermo Scientific) and introduced into mice (100 μg) via intraperitoneal injection. Mice were boosted with same dose of antigen at the indicated days along with Alum. Mice were also immunized with 25 μg NP₄₀-Ficoll (Biosearch Technologies) via intraperitoneal injection. Serum NP specific Ig levels in these mice were analyzed by ELISA. Briefly, 96 well ELISA plates (Nunc) were coated with NP₃₀-BSA (Biosearch Technology) overnight at 4°C, washed and blocked with 1% BSA fraction V (Sigma-Aldrich), serum titers were then added to the plates and incubated 4 h at 4°C. After washing alkaline phosphatase-labeled goat anti-mouse Ig isotype specific antibodies were added for 2 h at room temperature (SouthernBiotech). After washing, PNPP one component substrate (SouthernBiotech) was used to detect the amount of secondary antibody bound.

Boularan C., Hwang I.Y., Kamenyeva O., Park C., Harrison K., Huang Z, & Kehrl J.H. (2015). B-lymphocyte Specific loss of Ric-8A Results in a Gα Protein Deficit and Severe Humoral Immunodeficiency. Journal of immunology (Baltimore, Md. : 1950), 195(5), 2090-2102.

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Quantifying Antibody Responses to Immunogens

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WT and mutant mice were immunized with either sheep RBCs or TNP-KLH. For the sheep RBC immunizations 2 × 10⁸ sheep RBCs (Lonza Walkerville, Inc.) were given by intraperitoneal injection. TNP-KLH (Biosearch Technology) was mixed with Imject® Alum (Thermo Scientific) and introduced into mice (100 µg) via intraperitoneal injection. Mice were boosted with same dose of antigen at the indicated days along with Alum. Serum TNP specific Ig levels in these mice were analyzed by ELISA. Briefly, 96 well ELISA plates (Nunc) were coated with TNP-BSA (Biosearch Technology) overnight at 4°C, washed and blocked with 1% BSA fraction V (Sigma-Aldrich), serum titers were then added to the plates and incubated 4h at 4°C. After washing alkaline phosphatase-labeled goat anti-mouse Ig isotype specific antibodies were added for 2h at RT (SouthernBiotech). After washing, PNPP one component substrate (SouthernBiotech) was used to detect the amount of secondary antibody bound.

Hwang I.Y., Park C., Harrison K., Boularan C., Galés C, & Kehrl J.H. (2015). An Essential Role for RGS Protein/Gαi2 Interactions in B Lymphocyte Directed Cell Migration and Trafficking. Journal of immunology (Baltimore, Md. : 1950), 194(5), 2128-2139.

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