Cells were seeded and incubated for 24 h. Following this, the medium was replaced with fresh medium supplemented with TS. TS was added to the culture medium at a final concentration of 20 μmol/L for KYSE30, 60 μmol/L for HCT116, and 25 μmol/L for MC38. The cells were incubated further for 24 h, washed once, and immersed in PBS, and in vitro PDT was administered. The cells were further incubated for 15 min and the cell-surface expression levels of CRT (1:350 dilution; ab2907; Abcam, Cambridge, UK) and HSP90 (1:1,000 dilution; ab203126; Abcam) were analyzed by flow cytometry. The secondary antibody used was Alexa Fluor 488 goat anti-rabbit IgG (Thermo Fisher Scientific). All flow cytometry examinations were performed in triplicate on the FACSCanto II counting 10,000 events and analyzed using the FlowJo software.
Ab203126
Manufactured by Abcam
Sourced in United States
Ab203126 is a lab equipment product. It serves as a core function in scientific research and laboratory operations.
Automatically generated - may contain errors
Lab products found in correlation
Ab62631, by Abcam (2 mentions)
Ab6319, by Abcam (2 mentions)
Ab85615, by Abcam (2 mentions)
Ab8226, by Abcam (2 mentions)
Ecl kit, by Boster Bio (2 mentions)
Sc 33666, by Santa Cruz Biotechnology (2 mentions)
Ab6721, by Abcam (2 mentions)
Ab2907, by Abcam (1 mentions)
Alexa fluor 488 goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
Pannoramic midi, by 3DHISTECH (1 mentions)
Ab263899, by Abcam (1 mentions)
G box imaging system, by Syngene (1 mentions)
Biorad mini protean tetra system, by Bio-Rad (1 mentions)
Cw0102, by CWBIO (1 mentions)
Cw0103, by CWBIO (1 mentions)
Ab172730, by Abcam (1 mentions)
Ab1416, by Abcam (1 mentions)
Ab215715, by Abcam (1 mentions)
D085075, by Bio-Rad (1 mentions)
Ripa buffer, by Solarbio (1 mentions)
8 protocols using ab203126
1
Photodynamic Therapy Induces Immunogenic Cell Death
2
Uterine Tissue Immunolabeling Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The 4 μm sections of uterine tissues were used for immunolabeling analysis. Briefly, the uterine sections were permeabilized, blocked, and then incubated with primary antibodies against HSP90 (Cat. No. ab203126, 1:10,000, Abcam, USA), NLRP3 (Cat. No. ab263899, 1:2000, Abcam, USA), overnight at 4°C, followed by incubation with a corresponding secondary antibody at ordinary temperature for 1 h. Staining was developed using peroxidase 3,30-diaminobenzidine (DAB) substrate and counterstained with hematoxylin. Images were obtained using an Immunohistochemical scanner (Pannoramic MIDI, 3D histech, Hungary). The relative fluorescence intensity of positive cells was quantified using Image-Pro Plus software. The integrated optical density (IOD) and mean optical density (AOD) were quantified using Image-Pro Plus 6.0 software. AOD = IOD sum/area sum.
3
Western Blot Analysis of SOX9 and HSP90
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were lysed in EBC buffer and 2×SDS loading buffer to collect proteins. The protein samples were boiled at 100°C for 5 min and then electrophoresed in the Bio-Rad Mini PROTEAN Tetra System (Bio-Rad, USA). After electrophoresis, the proteins were transferred to the PVDF membrane under ice bath conditions, and then the membrane was washed twice with 1×TBST. The membrane was blocked with 5% free-fat milk for 2 h at room temperature. The primary antibodies to SOX9 (67439-1-Ig, Proteintech) and HSP90 (ab203126, Abcam) were diluted with 2% free-fat milk at ratios of 1:4,000 and 1: 10,000, respectively, and then incubated overnight at 4°C. Membrane was washed thrice for 15 min and incubated the secondary antibodies for 2 h at room temperature. After another three times washing, the bands were solarized and imaged using the Syngene G: BOX Imaging System (Cambridge, UK) (19 (link), 31 (link)).
4
Quantification of Target Proteins by Immunoblotting
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The levels of the target proteins were measured with the following primary antibodies: anti-CNPase (1:1,000, ab6319, Abcam), anti-myelin basic protein (MBP; 1:1,000, ab62631, Abcam), anti-neuron glia 2 (NG2; 1:1,000, sc-33666, SANTA CRUZ), anti-GSK3β (1:1,000, D5C5Z, CST), anti-phosphorylated (Ser9) GSK3β (p-ser9-GSK3β; 1:1,000, D85E12, CST), anti-5-HT1AR (1:1,000, ab85615, Abcam), anti-human gene and protein symbol ACTB/ACTB (β-actin; 1:5,000, ab8226, Abcam), and anti-heat-shock protein 90 (HSP90; 1:2,000, ab203126, Abcam). The corresponding secondary antibodies (goat anti-rabbit IgG, CW0103 and goat anti-rabbit IgG, CW0102, CWBIO, P. R. China) for each protein were applied at a dilution of 1:1,000, followed by development with an ECL kit (AR1111, Boster, P. R. China). The band intensity was quantified using Image Lab software (version 5.2.1).
5
Immunoprecipitation and Western Blot Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The cells were lysed in a pre-chilled RIPA lysis buffer, and the lysates were centrifuged at 14,000 g, 4°C for 15 min. The supernatant was collected and placed into another centrifuge tube. The tube was added with 500 μL of diluted rabbit or mouse antibodies against HSP90 (ab203126, 1: 100), E-cad (ab1416, 1: 100), or IgG (ab172730, 1: 20) (Abcam, Cambridge, MA, USA), and placed on a shaking bed at 4°C overnight or gently shaken at room temperature for 2 h. The tube was added with 100 μL of 50% protein A/G-agarose beads to capture antigen-antibody complexes and then shaken on a shaking bed at 4°C overnight or maintained at room temperature for 1 h. The mixture was centrifuged at 14,000 rpm for 5 s and the supernatant was removed. The antigen-antibody complexes were washed with 800 μL of pre-chilled RIPA buffer 3 times and resuspended in 60 μL of 2 × loading buffer. The loading samples were boiled for 5 min and then centrifuged at 14,000 g. The supernatant was boiled for 5 min before electrophoresis. Protein expression was detected by western blotting.
6
Protein Extraction and Western Blot Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Proteins from frozen kidney and heart tissues were collected using RIPA buffer (#R0010, Solarbio). Proteins (30 μg) were separated by 12% SDS PAGE (Bio-Rad, China) and transferred to PVDF membranes after being lysed. Before incubation with antibodies, membranes were cut and incubated with primary antibodies overnight at 4°C. Primary antibodies, including anti-rabbit HSP90 (1:500, #ab203126, Abcam), anti-rabbit TGF-β1 (1:500, #ab215715, Abcam) and anti-rabbit GAPDH (1:1000, #ab9487, Abcam), were diluted with 5% bovine serum albumin (BSA). After 3 times washing with TBS-0.01% Tween 20, the membranes were incubated with goat anti-rabbit IgG (HRP) (1:5000, #ab6721, Abcam) for 2 h at room temperature. After washing, the signals were visualized with an enhanced chemiluminescence reagent (#D085075, Bio-Rad). The proteins bands were quantified by ImageJ software and expressed as relative optical density.
7
Comprehensive Protein Analysis in Neuroscience
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The target proteins were measured with the following primary antibodies: anti-2'3'-cyclic nucleotide 3'phosphodiesterase (CNPase; mouse, 1:1000, ab6319, Abcam), anti-myelin basic protein (MBP; mouse, 1:1000, ab62631, Abcam), anti-NG2 (NG2; mouse, 1:1000, sc-33666, SANTA CRUZ), anti-SOX10 (SOX10; rabbit, 1:1000, ab155279, Abcam), anti-GSK3β (rabbit, 1:1000, D5C5Z, CST), anti-phosphorylated (Ser9) GSK3β (p-ser9-GSK3β; rabbit, 1:1000, D85E12, CST), anti-5-HT1AR (rabbit, 1:1000, ab85615, Abcam), antileucine-rich repeat and Ig domain containing 1 gene (LINGO1; rabbit, 1:1000, ab23631, Abcam), antimyelin-associated glycoprotein (MAG; mouse, 1:1000, ab89780, Abcam), anti-myelin oligodendrocyte glycoprotein (MOG; rabbit, 1:1000, ab233549, Abcam), anti-glyceraldehyde 3-phosphate dehydrogenase (GAPDH; rabbit, 1:5000, ab8245, Abcam), anti-human gene and protein symbol ACTB/ACTB (β-actin; mouse, 1:5000, ab8226, Abcam) and anti-heat-shock protein 90 (HSP90; rabbit, 1:2000, ab203126, Abcam). The corresponding secondary antibodies for each protein were used at a dilution of 1:1000, followed by development with an ECL kit (AR1111, Boster, China).
8
Western Blot Analysis of Protein Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Lysates from cells and tumor tissues were prepared and protein determined using the BCA assay (Beyotime Biotechnology). Protein (30 µg) from each sample was resolved on SDS-PAGE, and transferred to PVDF membrane. Membranes were blocked with 5% bovine serum albumin in Tris-buffered saline containing 0.1% Tween 20 and incubated with primary antibodies overnight at 4°C. Membranes were washed thrice, 5 min each time, and then incubated with either horseradish peroxidase (HRP)-conjugated goat anti-mouse (ab6789, Abcam, Cambridge, UK) or HRP-conjugated goat anti-rabbit (ab6721, Abcam) secondary antibodies for 2 h at room temperature. Immunoreactive bands were visualized using Pierce ECL plus Western blotting substrate (Thermo Fisher Scienti c, Waltham, MA, USA). The primary antibodies used were: ERα (sc-8002, Santa Cruz Biotechnology, Dallas, TX, USA), HSP90 (ab203126, Abcam), β-actin (4970S, Cell Signaling Technology, Boston, MA, USA), Bax (2774S, Cell Signaling Technology), Bcl-2 (15071S, Cell Signaling Technology), ubiquitin (3936S, Cell Signaling Technology), CYP1A1 (ab124295, Abcam) and AhR (ab190797, Abcam). The protein bands were analyzed using ImageQuant 5.2 software. β-actin was used as loading control.
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!