Methods of Protein Iodination
Erwin Regoeczi in Iodine-Labeled Plasma Proteins, 2019
Horseradish peroxidase catalyzes the oxidation of I- in a pH-dependent manner, optimal rates being observed at low pH (2.7 to 3.6).55,99 With less than stoichiometric quantities of peroxide, an equilibrium mixture of I-, I2, and is obtained.55 Ljunggren100 found no evidence for the enzyme to catalyze the iodination of free tyrosine at pH 7.4, but Pommier et al.101 obtained a good yield of iodotyrosines when working at pH 4.8. Bayse and Morrison55 compared the iodotyrosine-forming activities of horseradish peroxidase and lactoperoxidase, and found that on an equimolar basis, the activity of the former is about 14% of the latter.
Mitogenic Substances of Bupleuri Radix
Sheng-Li Pan in Bupleurum Species, 2006
To prove the presence of lignin, we studied the peroxidases (PODs) that are closely related to the synthesis and degradation of lignin. In general, Bupleuri radix and other crude drugs of plant origin are dried under appropriate conditions before use. It is believed that many plant PODs are more stable than other enzymes and their activities are retained after collection (Kolattukudy et al., 1992; Nicolas et al., 1994; Fujiyama et al., 1995; Wojtaszek, 1997; Piontek et al., 2001; Gajhede, 2001). One such generally used enzyme is horseradish peroxidase (HRP), which is essential for various immunochemical analyses.
Rigid monoclonal antibodies improve detection of SARS-CoV-2 nucleocapsid protein
Published in mAbs, 2021
Curtis D. Hodge, Daniel. J. Rosenberg, Patricia Grob, Mateusz Wilamowski, Andrzej Joachimiak, Greg L. Hura, Michal Hammel
To compare the relative affinity of each mAb for antigen, we performed binding kinetic assays. In addition, we performed assays on horseradish peroxidase (HRP)-conjugated mAbs in preparation for ELISAs, described below. Due to the high affinities of the mAbs, we opted to use a kinetic titration (single-cycle kinetics) strategy and avoid problematic regeneration steps (Materials and Methods). We measured the binding kinetics of mAb 1, 1-HRP, 2, 4, and 4-HRP by surface plasmon resonance (SPR) (Supplementary Figure 5). All antibodies (unconjugated and HRP-conjugated) had high-affinity constants (KD) in the picomolar range (Supplementary Table 1). The KD of HRP-conjugated mAb1 and 4 are very similar, at 11 and 28 pM, respectively. The percentage activity of the HRP-conjugated antibodies is lower than unconjugated, suggesting that conjugating HRP on the antibodies affects the percentage of available antibodies for interaction on the SPR sensor chip. The possibility exists that this effect could also be present in the chip-free solution-based ELISA. However, the high concentration of HRP-conjugated antibodies used (0.4 mg/mL; Methods), relative to the picomolar affinities, represents a large excess of functional, high-affinity HRP-conjugated antibodies in the ELISA. Therefore, the antibodies have comparable kinetics, effectively excluding them as explanations for functional outcomes.
Fabrication and transplantation of chitosan-selenium biodegradable nanocomposite conduit on transected sciatic nerve: a novel study in rat model
Published in Neurological Research, 2020
Salar Dolkhani, Alireza Najafpour, Rahim Mohammadi
In this study, anti-S-100 (1:200, DAKO, USA) was used as marker for myelin sheath. Specimens were post fixed with 4% paraformaldehyde for 2 h and embedded in paraffin. Prior to immunohistochemistry nerve sections were dewaxed and rehydrated in PBS (pH 7.4). Then, the nerve sections were incubated with 0.6% hydrogen peroxide for 30 min. To block non-specific immunoreactions the sections were incubated with normal swine serum (1:50, DAKO, USA). Sections were then incubated in S-100 protein antibody solution for 1 h at room temperature. They were washed three times with PBS and incubated in biotinylated anti-mouse rabbit IgG solution for 1 h. Horseradish peroxidase-labelled secondary antibody was applied for 1 h. After that all sections were incubated with 3,3ʹ- diaminobenzidine tetrahydrochloride chromogene substrate solution (DAB, DAKO, USA) for 10 min. The results of immunohistochemistry were examined under a light microscope.
Assessments of regenerative potential of silymarin nanoparticles loaded into chitosan conduit on peripheral nerve regeneration: a transected sciatic nerve model in rat
Published in Neurological Research, 2021
Pouria Ebrahimi-Zadehlou, Alireza Najafpour, Rahim Mohammadi
In this study, anti-S-100 was used as marker for myelin sheath. Specimens were post fixed with 4% paraformaldehyde for 2 h and embedded in paraffin. Prior to immunohistochemistry nerve sections were dewaxed and rehydrated in PBS (pH 7.4). Then the nerve sections were incubated with 0.6% hydrogen peroxide for 30 minutes. To block non-specific immunoreactions the sections were incubated with normal swine serum. Sections were then incubated in S-100 protein antibody solution for 1 h at room temperature. They were washed three times with PBS and incubated in biotinylated anti-mouse rabbit IgG solution for 1 h. Horseradish peroxidase-labeled secondary antibody was applied for 1 h. After that all sections were incubated with 3,3ʹ- diaminobenzidine tetrahydrochloride chromogene substrate solution for 10 min. The results of immunohistochemistry were examined under a light microscope.
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