High binding plates were washed with Na2CO3 buffer 3 times and coated with 50 μL of a 1:1000 dilution of anti-ApoA-I in Na2CO3 buffer for 1 hour at 37 °C. Plates were washed 6 times with PBS-T and blocked with PBS containing 1% BSA and 0.1% Tween (PBS-B) for 1 hour at 37 °C prior to 2 washes with PBS-T. Samples were diluted 1:100,000 in PBS-B and 100 μL of diluted plasma was added to each well and incubated for 30 min at 37 °C. Following 6 washes with PBS-T, 100 μL of anti-ApoA-I (Millipore Sigma, 178422), diluted 1:2000 in PBS-B, was added and incubated for 30 min at 37 °C. After 6 washes with PBS-T, 100 μL of anti-rabbit IgG HRP (abcam, ab6721) diluted 1:20,000 in PBS-B was added and incubated for 30 min at 37 °C. The plates were then washed, developed with TMB, quenched with H2SO4 and the absorbance measured at 450 nm. Samples were analyzed in duplicate with a standard curve on each plate using purified human ApoA-I (Athens Research and Technology 16-16-120101).
Anti rabbit igg hrp
Manufactured by Abcam
Sourced in United Kingdom, United States
Anti-rabbit IgG-HRP is a secondary antibody conjugated with horseradish peroxidase (HRP). It is designed to bind to and detect rabbit primary antibodies in various immunoassays and immunochemical techniques.
Automatically generated - may contain errors
Lab products found in correlation
Anti mouse igg hrp, by Abcam (3 mentions)
Nitrocellulose membrane, by Cytiva (2 mentions)
Protease inhibitor cocktail, by Roche (2 mentions)
Nitrocellulose membrane, by GE Healthcare (1 mentions)
Westernbright quantum, by Advansta (1 mentions)
Duoset elisa kit, by R&D Systems (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Ecl kit, by Merck Group (1 mentions)
Cell lysis reagent, by Merck Group (1 mentions)
Rabbit anti β actin polyclonal antibody, by Santa Cruz Biotechnology (1 mentions)
Anti neun antibody, by Abcam (1 mentions)
Vector vip substrate, by Vector Laboratories (1 mentions)
Immpact dab substrate, by Vector Laboratories (1 mentions)
Ecl western blotting substrate, by Thermo Fisher Scientific (1 mentions)
Anti flag hrp, by Merck Group (1 mentions)
Ecl detection kit, by Thermo Fisher Scientific (1 mentions)
Metformin, by Merck Group (1 mentions)
Snap i d protein detection system, by Merck Group (1 mentions)
Anti gapdh, by Cell Signaling Technology (1 mentions)
Il 1β, by R&D Systems (1 mentions)
20 protocols using anti rabbit igg hrp
1
Quantitative ApoA-I Immunoassay Protocol
2
Inflammasome Activation and IL-1β Analysis
3
Protein Extraction and Western Blot Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After treatment, cells were washed twice with cold PBS and then pelleted, and then cell lysis reagent (Sigma-Aldrich) was added to the cell pellet for protein extraction. Protein concentration was determined by bovine serum albumin (BSA) assay using BSA as the reference, and the absorbance of the reacted product was measured at wavelength of 540–595 nm. Samples were boiled for 3 min before loading onto 12% sodium dodecyl sulfate (SDS)-polyacrylamide gel. After electrophoresis, the gels were electro blotted onto a polyvinylidene fluoride (PVDF) membranes (Merck Millipore, Whitehouse Station, NJ, USA). The antibodies used were as follows: anti-Bcl-2 rabbit polyclonal antibody (Abcam, Cambridge, MA, USA), anti-Bax rabbit polyclonal antibody (Abcam), anti-β-actin rabbit polyclonal antibody (Santa Cruz Biotechnology Inc., Dallas, TX, USA), and anti-rabbit IgG HRP (Abcam). The blot was visualized with an enhanced chemiluminescence (ECL) kit (Merck Millipore) and exposed to ECL Hyperfilm.
4
Dual Immunohistochemical Staining of Mouse and Human Brain Tissues
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Mouse brain sections were incubated with anti-mouse transferrin (primary antibody; Bethyl) followed by anti-goat IgG-HRP (secondary antibody; Jackson Immuno Research Inc., West Grove, PA, USA). Human brain sections were incubated with anti-human transferrin antibody (DakoCytomation, Glosyrup, Denmark) followed by HRP-conjugated secondary antibody (ab205718, abcam). Signals were developed with ImmPACT DAB substrate (Vector Lab.). For double staining, DAB-stained slides were treated with quenching peroxidase and then with anti-NeuN antibody (abcam). The sections were further incubated with anti-rabbit IgG-HRP (abcam). Signals were developed with VECTOR VIP substrate (Vector Lab.).
5
Immunoblotting Analysis of Transfected Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For immunoblotting analysis, cells were transfected as for the reporter assays, but scaled twofold onto a 12 well, and replacing the Cas9/sgRNA plasmids with EV. Subsequently, cells were extracted using NETN buffer (20 mM Tris at pH 8.0, 100 mM NaCl, 1 mM EDTA, 0.5% Igepal, 1.25 mM DTT, Roche (Basel, Switzerland) protease inhibitor) with several freeze/thaw cycles. Extracts were probed with antibodies for mouse monoclonal anti-FLAG HRP (Sigma-Aldrich Cat#A8592), or rabbit polyclonal anti-ACTIN (Sigma-Aldrich (MiliporeSigma) Cat#A2066) with the secondary antibody goat polyclonal Anti-Rabbit IgG HRP (Abcam, Cambridge, U.K. Cat#ab205718). ECL western blotting substrate (ThermoFisher Cat#32106) was used to develop HRP signals.
6
Metformin Modulates AMPK and Autophagy in Chondrocytes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Chondrocytes (1 × 106 cell/well) were incubated with IL-1β 20 ng/mL (R&D Systems, MN, USA) and metformin 0.2, 1, and 5 mM (Sigma Aldrich) for 1 h. Proteins were separated by SDS-PAGE and transferred onto a nitrocellulose membrane (Amersham Pharmacia Biotech, Piscataway, NJ, USA). Western blotting was performed using a SNAP i.d. protein detection system (Millipore). The hybridized bands were detected using an enhanced chemiluminescence (ECL) detection kit (Thermo Fisher Scientific, MA. USA) and the following antibodies: anti-phospho-AMPK (1:1000 dilution), anti-AMPK (1:1000 dilution, Cat. No. #2535), anti-LC3B (1:2000 dilution, Cat. No. #2775), anti-p62 (1:400 dilution, Cat. No. #5114), anti-caspase-1 (1:1000 dilution, Cat. No. #2225), anti-caspase-3 (1:1000 dilution, Cat. No. #9662) (all from Cell Signaling Technologies), anti-GAPDH (1:2000 dilution, Cat. No. ab9485, Abcam), and anti-rabbit IgG-HRP (1:2000 dilution, Cat. No. sc2357, Santa Cruz Biotechnology, Dallas, Texas, USA). The Western blot bands were quantified using the Fiji/ImageJ program.
7
Protein Expression Analysis in Cells
8
Phosphorylation Dynamics in MOVAS Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
MOVAS-1 cell lysates from 1 h, 12 h, and 24 h old cultures (30 µg of protein) were separated on a 12% polyacrylamide gel under reducing conditions and transferred onto a nitrocellulose membrane (Hybond-ECL, Amersham, Mississauga, Ontario, Canada). The membrane was blocked with 5% non-fat milk in PBS and probed with either rabbit anti-phospho-ERK1/2 (1:1000, Abcam), rabbit anti-ERK1/2 (1:1000, Abcam), or rabbit anti-β actin (1:5000, ThermoFisher, Burlington, Ontario, Canada) antibodies in PBS, 5% non-fat milk, and 0.1% Tween 20 overnight at 4 °C. Anti-rabbit IgG-HRP (Abcam) was used as a secondary antibody and membranes were incubated for 1 h at room temperature. Finally, membranes were developed by chemiluminescence (Amersham).
9
Bufalin-Induced Apoptosis and Stemness Regulation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Bufalin was purchased from Sigma-Aldrich (St. Louis, MO, USA). The antibody against glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was purchased from PeproTech (Rocky Hill, NJ, USA). The following antibodies were obtained from Cell Signaling Technology (Boston, MA, USA): β-actin rabbit mAb [horseradish peroxidase (HRP) conjugated], caspase 3, Bcl-2, Bcl-xL, caspase 9, and poly(ADP-ribose) polymerase (PARP). The antibodies against Bad and p-Bad were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). The antibodies against SOX2, Nanog, Oct4, and hTERT anti-mouse IgG-HRP and anti-rabbit IgG-HRP were purchased from Abcam (San Antonio, TX, USA). RNAiso Plus kits, PrimeScript Reverse Transcriptase kits, and SYBR Premix Ex Taq II kits were purchased from Takara (Tokyo, Japan). The hTERT and β-actin gene primers were synthesized by Sangon Biotech Co. Ltd. (Shanghai, P.R. China). Recombinant human epidermal growth factor (EGF; Invitrogen, Carlsbad, CA, USA), basic fibroblast growth factor (bFGF; EMD Millipore, Billerica, MA, USA), growth factor B-27 (Gibco, Grand Island, NY, USA), growth factor N2 (Gibco), and heparin (Selleck, Houston, TX, USA) were used for spheroid culture.
10
Kidney Tissue Protein Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The preparation of kidney tissue homogenates and western blot analysis of protein expression were performed using routine procedures as described previously22 (link). The following antibodies were used: rabbit anti-rat Wnt4 (1:200, Santa Cruz Biotechnology, USA), rabbit anti-rat β-catenin (1:5000, Abcam, UK), and anti-rabbit IgG HRP (1:5000; Abcam, UK). All of the western blot results were normalized to β-actin.
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!