The largest database of trusted experimental protocols

7 protocols using streptavidin peroxidase detection system

1

Immunohistochemical and Immunofluorescent Analyses

Check if the same lab product or an alternative is used in the 5 most similar protocols
For immunohistochemistry staining, formalin-fixed and paraffin-embedded tissue sections were incubated with primary antibodies against MTCO1 (ab14705, Abcam), CD68 (ab955, Abcam), or CD3 (ab16669, Abcam) and then analyzed using streptavidin peroxidase detection system (Maixin) according to the manufacturer’s protocol. Diaminobenzidine (DAB) (Maixin) was used as a horseradish peroxidase (HRP)–specific substrate. For immunofluorescence staining, formaldehyde-fixed cells or kidney sections were performed with primary antibodies against RFP (ab62341, Abcam), CD63 (sc5275, Santa Cruz Biotechnology), IL-10 (ab9969, Abcam), KIM-1 (MA5-28211, Invitrogen), CD68 (ab955, Abcam), iNOS (ab15323, Abcam), CD206 (ab64693, Abcam), and CD3 (ab16669, Abcam), followed by incubation with secondary antibodies. Cell nuclei were stained with DAPI. Immunostained samples were visualized under a confocal microscope (FV1000, Olympus).
+ Open protocol
+ Expand
2

Immune Cell Profiling in Kidney Tissue

Check if the same lab product or an alternative is used in the 5 most similar protocols
For IHC, formalin-fixed and paraffin-embedded kidney sections were incubated with primary antibodies against CD68 (ab955, Abcam), neutrophil (ab2557, Abcam), or CD3 (ab16669, Abcam), and then analyzed using streptavidin peroxidase detection system (Maixin) according to the manufacturer's protocol. DAB (Maixin) was used as an HRP-specific substrate. Quantitative analysis of the number of positive cells was performed under original magnification (×400) in 40 randomly selected fields per mice in a blinded fashion. Immunofluorescence staining of formaldehyde-fixed cells was performed with primary antibody against NF-κB p-p65 (3033, CST), followed by incubation with a secondary antibody (Invitrogen, USA). Cell nuclei were stained with DAPI. Immunostained samples were visualized under a confocal microscope. Images were quantified by counting the number of positive nuclei and dividing by the total number of nuclei.
+ Open protocol
+ Expand
3

Renal Tissue Analysis: PAS Staining and Immunohistochemistry

Check if the same lab product or an alternative is used in the 5 most similar protocols
Renal tissues were used for periodic acid-Schiff (PAS) staining. The tubular injury was semiquantitatively scored as follows 28 (link): 0, no damage; 1, <25%; 2, 25~50%; 3, 50~75%; 4, >75%. The tubular injury score was calculated as the average score from 10 random sections. For immunohistochemistry staining, 4-μm-thick renal tissue sections were performed with primary antibodies against CD68 (ab955, Abcam), or CD3 (ab16669, Abcam) and analyzed using streptavidin peroxidase detection system (Maixin) as instructing by the protocols. For immunofluorescence analysis, tissue sections were incubated with primary antibodies against kidney injury molecular-1 (KIM-1; MA5-28211, Invitrogen), Ki-67(ab15580, Abcam), and p-H3 (ab14955, Abcam), followed by incubation with secondary antibodies (ab150114 and ab150077, Abcam). The proximal tubules were labeled by Lotus tetragonolobus lectin (LTL, FL-1321, vector labs) 29 (link),30 (link). The number of positive tubules and cells was counted under a confocal microscope in ten random fields per mouse in a blinded manner.
+ Open protocol
+ Expand
4

Immunohistochemical Analysis of Cx43 and Podocin

Check if the same lab product or an alternative is used in the 5 most similar protocols
Paraffin-embedded sections were used for immunohistochemistry. Briefly, the sections were incubated with primary antibodies against Cx43 and podocin overnight at 4°C. The sections were then analyzed using a streptavidin peroxidase detection system (Maixin) according to the manufacturer's protocol. Reactions were conducted using a DAB substrate kit (Maixin) and counterstaining was performed using hematoxylin. The sections were visualized under a microscope (Nikon Corporation).
+ Open protocol
+ Expand
5

Immunohistochemical Evaluation of Kidney Injury Markers

Check if the same lab product or an alternative is used in the 5 most similar protocols
For immunohistochemistry staining, after antigen retrieval using the antigen retrieval buffer EDTA at pH 9.0 and heating for 15 min at 100 °C, the formalin-fixed and paraffin-embedded tissue sections of kidney were incubated with primary antibodies against kidney injury molecular-1 (KIM-1, MA5–28211, Invitrogen), F4/80 (ab6640, Abcam, Cambridge, MA), HK2 (ab209847, Abcam), PTEN-induced putative kinase 1 (PINK1, ab23707, Abcam), Bcl-2 19-kDa interacting protein 3 (BNIP3, ab109362, Abcam), or cytochrome C oxidase subunit I (ab14705, Abcam), and thereafter analyzed using a streptavidin peroxidase detection system (Maixin Biotech, Fuzhou, China) in accordance with the manufacturer’s protocol. Diaminobenzidine (Maixin) was used as horseradish peroxidase (HRP)-specific substrate.
+ Open protocol
+ Expand
6

Immunohistochemistry and Immunofluorescence Staining of IgA in Intestine

Check if the same lab product or an alternative is used in the 5 most similar protocols
For IHC staining, formalin-fixed and paraffin-embedded small intestine sections were incubated with primary antibodies against IgA (Abcam, ab97234) and analyzed using streptavidin peroxidase detection system (Maixin) according to the manufacturer’s protocol. DAB (Maixin) was used as an HRP-specific substrate. Staining intensity for IgA was ranked using a 5-point scale from 0 (unstained) to 4 (very intensively stained). The histopathological evaluations were performed by three independent pathologists. Every group and tissue examination were blinded.
The kidney and small intestine tissue for IF staining was cut into frozen sections and fixed with acetone for 1 min. After fixation, they were blocked with 2% bovine serum albumin diluted by PBS at room temperature for 1 h. They were washed with PBS three times and incubated with FITC-labelled goat anti-mouse IgA (Abcam, ab97234) at 37°C for 40min. After washing with PBS three times, they were mounted with anti-quenching tablets and observed under a confocal microscope. Cell nuclei were stained with DAPI. Images were quantified by counting the number of positive nuclei and divided by the total number of nuclei.
+ Open protocol
+ Expand
7

Immunohistochemical Analysis of NLRP3 Inflammasome

Check if the same lab product or an alternative is used in the 5 most similar protocols
Paraffin-embedded sections from the kidney cortex were used for immunohistochemistry. Briefly, the sections were incubated with primary antibodies to F4/80, neutrophil (Abcam, USA), NLPR3 (Adipogen, USA), ASC, caspase-1 (Santa Cruz Biotechnology, USA) overnight at 4 °C. The sections were then analyzed using streptavidin peroxidase detection system (Maixin, China) according to the manufacturer’s protocol. The reaction was developed using DAB substrate kit (Maixin, China), and counterstaining was performed using hematoxylin.
For analysis and localizing the expression of the NLRP3 and cathepsin B, immunofluorescence staining of tissue sections or formaldehyde-fixed cells was performed using anti-NLRP3, anti-cathepsin B antibodies (Abcam, USA), respectively in a humidified chamber overnight at 4 °C, followed by incubation with an Alexa fluorescein-labeled secondary antibodies (Invitrogen, USA) for 1 h. Cell nuclei were stained with DAPI. Immunostained samples were visualized under a confocal microscope. Immunofluorescence for p65 (Cell Signaling Technology, USA) was similarly performed. Quantification of intensity of p65 in nuclear was performed by measuring area, integrated density, and mean gray value using Image-Pro Plus.
+ Open protocol
+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required

Sign up now

Revolutionizing how scientists
search and build protocols!