The largest database of trusted experimental protocols

4 protocols using anti apoa1

1

Extracellular Vesicle Protein Profiling

Check if the same lab product or an alternative is used in the 5 most similar protocols
Proteins (12 μg) from each group were separated by SDS–polyacrylamide gel electrophoresis and then transferred onto polyvinylidene difluoride (PVDF) membranes. PVDF membranes were blocked for 1 hour at room temperature and then incubated with primary antibody overnight at 4°C. The following antibodies were used for Western blot analysis: anti-CD63 (1:1000; Abcam, ab134045), anti-TSG101 (1:1000; Abcam, ab125011), anti-Alix (1:1000; Abcam, ab186429), anti–Flotillin-2 (1:1000; Abcam, ab181988), anti-ARF6 (1:1000; Abcam, ab13126), anti-APOA1 (1:1000; Abcam, ab52945), anti-APOA2 (1:1000; Abcam, ab92478), anti-APOB (1:1000; Abcam, ab139401), and anti-ALB (1:1000; Abcam, ab207327). Horseradish peroxidase–conjugated goat anti-rabbit and goat anti-mouse antibodies were used to detect the bound primary antibodies. The signals were detected by the enhanced chemiluminescence reagent. Data from the bands were determined through ImageJ software.
+ Open protocol
+ Expand
2

Antibody Panel for Extracellular Vesicle Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols
The following antibodies (Abs) were used in the experiments of this study: anti‐CD9 (BioLegend, Cat# 312102; San Diego, CA, USA), anti‐CD63 (Novus Biologicals, Cat# NBP2‐42225; Centennial, CO, USA), anti‐CD81 (BioLegend, Cat# 349502), anti‐IGF1R (ganitumab, a kind gift from Dr. J.C. Williams), anti‐cytochrome C (BD Biosciences, Cat# 556432; Franklin Lakes, NJ, USA), anti‐syntenin (Abcam, Cat# ab19903; Waltham, MA, USA) and anti‐ApoA1 (Abcam, Cat# ab52945) Abs, as well as goat anti‐rabbit (anti‐rabbit IgG; Invitrogen, Cat# A16112; Waltham, MA, USA), IRDye 800CW goat anti‐mouse (Li‐Cor Biotechnology, Cat# 926–32210; Lincoln, NE, USA) and IRDye 680RD goat anti‐rabbit (Li‐Cor Biotechnology, Cat# 926–68071) secondary Abs. For ExoView experiments, fluorescently labelled anti‐CD9‐CF488A, anti‐CD63‐CF647 and anti‐CD81‐CF555 Abs from the manufacturer's kit were used (details below). Human lactadherin (MFG‐E8), aa Leu24‐Cys387 (contains both C1 and C2 domains) was purchased from Novus Biologicals (Cat# 2767‐MF‐050).
+ Open protocol
+ Expand
3

Western Blot Analysis of Extracellular Vesicles

Check if the same lab product or an alternative is used in the 5 most similar protocols
For western blot (WB) analysis, the purified extracellular vesicles were lysed with radio-immunoprecipitation assay buffer (Santa Cruz Biotechnology, Dallas, TX, USA), and the cleared lysate was collected by centrifugation for protein separation on 12% sodium dodecyl sulfate-polyacrylamide (SDS-PAGE) gels. After electrophoresis, the separated proteins were transferred to 0.45 μm polyvinylidene difluoride (PVDF) membranes (Millipore, USA). The membranes were blocked for 1 h with Tris-buffered saline containing Tween 20 (TBST) with 5% non-fat milk. The blots were then incubated with primary antibody at 4°C overnight. The primary antibodies used included mouse monoclonal anti-CD63 (Abcam, Cambridge, UK), anti-ASFV p30 (Prepared by our laboratory), rabbit monoclonal anti-CD9 (Abcam, Cam bridge, UK), anti-APOA1(Abcam, Cam bridge, UK), rabbit polyclonal anti-ASFV p72 (prepared by our laboratory), SERPINC1 (Abcam, Cam bridge, UK). After washing three times with TBST, the membranes were incubated with horseradish peroxidase (HRP)-labeled secondary antibody (Proteintech, Chicago, IL, USA) for 2 h at room temperature. Finally, the proteins were visualized with Clarity enhanced chemiluminescence (ECL) WB substrate (Bio-Rad Laboratories, Hercules, CA, USA).
+ Open protocol
+ Expand
4

N-arachidonoyl glycine photocrosslinking probe

Check if the same lab product or an alternative is used in the 5 most similar protocols
N-arachidonoyl glycine was purchased from Cayman. N-acyl amino acid photocrosslinking probe was synthesized as previously described (Long et al., 2016 ). Phenylalanine (BP391100) and oleic acid (AC270290050) were purchased from Fisher. BSA (protease free, fatty acid free, essentially globulin free) was purchased from Sigma (A7030). Pooled human plasma (IPLA-N) was purchased from Innovative Research. The following antibodies were used: anti-Flag M2 (Sigma F3165), anti-APOB (Abcam ab20737), anti-APOA1 (ab7614). The following plasmids were used: mouse AAV-PM20D1-flag (Addgene 132682), mouse PM20D1-flag (Addgene 84566). Adeno-associated viruses were produced at the Penn Vector Core. AAV-GFP virus was purchased from Penn Vector Core (AAV8.CB7.CI.eGFP.WPRE.rBG).
+ 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!