Human retinal microvascular endothelial cells were seeded at a density of 2 × 104 cells/well in 24-well plates and counted in a cell number counter after 48 h of continuous treatment with endothelial cell growth media-2 or PDR vitreous (1:3 dilution in EGM-2). At least three independent experiments were performed as described previously (6 (link)). In addition, immunofluorescent staining for Ki67 (rabbit anti-Ki67; Proteintech, Rosemont, IL US) for marking proliferating cells was performed as previously described (17 (link)).
Rabbit anti ki67
Manufactured by Proteintech
Sourced in United States
Rabbit anti-Ki67 is a primary antibody that binds to the Ki67 protein. Ki67 is a nuclear protein expressed during all active phases of the cell cycle (G1, S, G2, and mitosis), but is absent in resting cells (G0). This antibody can be used to detect actively proliferating cells in various applications.
Automatically generated - may contain errors
Lab products found in correlation
5 protocols using rabbit anti ki67
1
Proliferation of Retinal Endothelial Cells
2
Immunofluorescence Staining of SACC Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The SACC cells were cultured in 8-chamber glass slide until the desired time points. Firstly, aspirated the upper medium and add fixative containing 4 % PFA, 5 % sucrose and 0.1 % Triton X-100 for 10 min. Secondly, aspirated the fixative and added PBS to each well to incubate for 10 min. Then, aspirated the PBS and add 0.05 % Tween 20 for 20 min. After 1 h blocking by 5 % goat serum, the mouse anti-DEC2 and the rabbit anti-ki67 (1:50; Proteintech) was added to the sections to incubate overnight at 4 °C. The next day, aspirated the antibody and washed each well with PBS third times. Mixed rhodamine red-conjugated anti-mouse IgG (H + L) (DyLight™680 Conjugate) and FITC-anti-mouse IgG (H + L) (DyLight™ 800 Conjugate) in 1:1 ratio. Incubated fixed cells with the mixed secondary antibodies for 1 h at 37 °C, then washed each well with PBS and counterstained the DAPI for 10 min. Images were obtained by a fluorescence microscope (Olympus BX51).
3
Immunofluorescence Staining of Ki67
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells cultured on slides or coverslips were fixed with 4% paraformaldehyde for 10 min at room temperature, followed by permeabilization with 0.3% Triton X-100 in PBS. Sections were incubated with a blocking buffer containing 5% BSA in PBS for 1 h at room temperature, followed by addition of rabbit anti-Ki67 (Proteintech) and incubated overnight at 4°C. Primary Abs were labeled with secondary Abs conjugated to the fluorescent probes, and nuclei were labeled with DAPI. Slides were covered with a coverslip with ProLong Gold antifade reagent (Invitrogen) and allowed to dry for 24 h at room temperature. Images were captured with a 20X objective.
4
Immunohistochemical Analysis of DEC2, Ki-67, and NR2F1
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Paraffin-embedded sections were cut into 4um and deparaffinized in xylene and rehydrated, and endogenous peroxidase was blocked with 3%H2O2. Antigen retrieval was accomplished by 0.01 mol/L citrate buffer solution (pH 6.0) in a 700 W microwave oven for 15 min. After incubation with 5% normal goat serum for 20 min, the slides were exposed overnight at 4 °C to the rabbit anti-DEC2 (1:150; Proteintech), rabbit anti-Ki-67(1:800; Proteintech), rabbit anti-NR2F1 (1:200; Proteintech). Sections were then incubated with biotinylated goat anti-rabbit IgG (Zhongshan Goldenbridge Biotechnology) for 1 h, and streptavidin-peroxidase for 30 min. The 0.02% diaminobenzidine tetrahydrochloride was used as a chromogen, and the slides were counterstained with hematoxylin. The percentage of positive cells was estimated using an image analysis system (Leica).
5
Immunohistochemical Staining of Paraffin-Embedded Tissues
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Paraffin-embedded tissues sections were deparaffinized and rehydrated with xylene, gradient ethanol (70, 80, 95, 100%) and incubated for 20 min in 3% H2O2 to block endogenous peroxidase activity. After blocking with 10% goat serum and then incubated with 50 μl rabbit anti-Ki67 (Proteintech, Wuhan, China), rabbit anti-MATR3 (Proteintech, Wuhan, China), rabbit anti-NCALD (Proteintech, Wuhan, China) at 4 °C overnight. Subsequently, sections were incubated with an appropriate secondary antibody for 20 min at 37 °C and developed with diaminobenzidine and stained with hematoxylin for 5 min. After each treatment, the sections were dehydrated, cleared, mounted, and viewed under the microscope.
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
Revolutionizing how scientists
search and build protocols!