Liver protein isolation and western blots on liver extracts and plasma samples were performed as previously described.17 (link) Primary antibodies to the 2A peptide (1:5000, rabbit, ABS31, Sigma-Aldrich), GFP (1:3000, rabbit, A-11122, Fisher-Scientific), FAH (1:1000, rabbit, SAB2100745, Sigma-Aldrich), apoA1 (1:5000, rabbit, K23500R, Meridian) and beta tubulin (1:500, mouse, University of Iowa Developmental Studies Hybridoma Bank E7) were diluted in 1% BSA in PBS-T and membranes were incubated overnight at 4°C. Goat anti-rabbit 680 nm and anti-mouse 800 nm antibodies (1:15000, 611-144-002-0.5 and 610-145-002-0.5, Rockland) were incubated at room temperature for 1 hour and imaged using an Odyssey Classic (Li-Cor). Red Ponceau stain was used as loading control for western blot on plasma samples.
Abs31
Manufactured by Merck Group
Sourced in United States, Italy
The ABS31 is a laboratory instrument designed for the detection and quantification of targeted analytes in various sample types. It utilizes advanced spectroscopic techniques to provide accurate and reliable results. The core function of the ABS31 is to enable precise measurement and analysis of samples, supporting research and diagnostic applications.
Automatically generated - may contain errors
Lab products found in correlation
Ecl prime, by Cytiva (2 mentions)
Chemidoc xrs system, by Bio-Rad (2 mentions)
A11122, by Thermo Fisher Scientific (1 mentions)
Odyssey classic, by LI COR (1 mentions)
Apoptag peroxidase in situ apoptosis detection kit, by Merck Group (1 mentions)
Ab151998, by Abcam (1 mentions)
Ab81289, by Abcam (1 mentions)
Rabbit on rodent hrp polymer rmr622h, by Biocare Medical (1 mentions)
Ci l bright field microscope, by Nikon (1 mentions)
Crm325, by Biocare Medical (1 mentions)
Iblot, by Thermo Fisher Scientific (1 mentions)
Ap156p, by Merck Group (1 mentions)
β actin, by Abcam (1 mentions)
Ab97046, by Abcam (1 mentions)
Ac 15, by Abcam (1 mentions)
Pab416, by Abcam (1 mentions)
Polyvinylidene fluoride (pvdf), by Merck Group (1 mentions)
Ab8227, by Abcam (1 mentions)
Hrp conjugated secondary antibody, by Agilent Technologies (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
7 protocols using abs31
1
Liver Protein Isolation and Western Blot Analysis
2
Immunohistochemical Analysis of Liver Tissue
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Livers were fixed in 10% formalin overnight at 4°C, then gradually dehydrated with ethanol and embedded in paraffin. Immunohistochemistry and hematoxylin and eosin (H&E) staining were performed by the Texas Digestive Diseases Morphology Core at Baylor College of Medicine, as previously described.17 (link) The following primary antibodies were used for immunohistochemistry: rabbit anti 2A peptide (1:7500, ABS31, Sigma-Aldrich), rabbit anti Ki67 (1:60, CRM325, Biocare); rabbit anti FAH (1:65, ab151998, Abcam); rabbit anti Cd34 (1:250, ab81289, Abcam). The Ki67, FAH and Cd34 antibodies were then detected with a Rabbit-on-Rodent HRP-Polymer (RMR622H, Biocare) and visualized with DAB chromogen (DB801, Biocare). The 2A antibody was detected and visualized using a Leica Bond Polymer Detection kit (DS9800). Tunel staining was performed and detected using ApopTag Peroxidase In Situ Apoptosis Detection Kit (Millipore, S7100). Reticulin staining was performed using the Epredia Reticulin Sliver Stain Kit (87025, Epredia), following the manufacturer instructions. Slides were counterstained with hematoxylin, dehydrated, and mounted with a permanent mounting medium. A Nikon Ci-L bright field microscope was used for imaging at the Integrated Microscopy core (Baylor College of Medicine).
3
Protein Extraction and Western Blotting
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Protein extraction and western blotting was performed using standard procedures. In brief, 30 μg protein per sample was separated by Bolt® Bis‐Tris 4–12% SDS/PAGE (Thermo Fisher) and blotted onto polyvinylidene fluoride (PVDF) by iBlot® (Invitrogen, Carlsbad, CA, USA) and blocked by 4% skim milk in 2% Tris‐buffered saline/Tween. Primary antibodies used in this study were as follows: 2A peptide 1 : 2000 ABS31 (Millipore, Billerica, MA, USA), SV40LT 1 : 400 Pab416 (Abcam, Cambridge, UK), and β‐actin 1 : 10 000 AC‐15 (Abcam). Secondary horseradish peroxidase (HRP) antibodies used in this study were as follows: goat anti‐rabbit (AP156P; Millipore) and rabbit anti‐mouse (ab97046; Abcam).
4
Immunoblotting of FMDV Proteins
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Immunoblotting was performed according to standard methods as described previously [26 (link)]. Briefly, samples were mixed with Laemmli sample buffer (with 25 mM dithiothreitol), separated by SDS-PAGE (12.5% or 15% polyacrylamide) and transferred to polyvinylidene difluoride membranes (PVDF, Millipore). These were incubated with primary antibodies specific for the FMDV capsid proteins (anti-FMDV O1 Manisa guinea pig serum), FMDV 2A (ABS31, Millipore), actin (ab8227, Abcam) or FMDV 3Cpro (anti-FMDV 3C 1G1, kindly provided by E. Brocchi, Brescia, Italy, as used previously [30 (link)]. Immunoreactive proteins were visualized using appropriate secondary horseradish peroxidase-conjugated antibodies (Dako) and a chemiluminescence detection kit (ECL Prime, Amersham) with a Chemi-Doc XRS system (Bio-Rad).
5
Immunoblotting FMDV VP2 and 2A Proteins
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Immunoblotting was performed, using cell lysates, according to standard methods as described previously [21 (link)]. Briefly, aliquots of cell lysate were mixed with Laemmli sample buffer (with 25 mM DTT), and the proteins were separated by SDS-PAGE (12.5 or 4–15 % polyacrylamide) and transferred to PVDF membranes (Millipore). Specific proteins were detected with primary antibodies recognizing FMDV VP2 (monoclonal antibody 4B2, a gift from L. Yu, as described by Yu et al. [32 (link)]) and FMDV 2A (ABS31; Millipore). Bound proteins were visualized using appropriate HRP-conjugated secondary antibodies (Dako) and a chemiluminescence detection kit (ECL Prime, Amersham) with a Chemi-Doc XRS system (Bio-Rad).
6
Ebola Virus Protein Detection
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Full length EBOV NP was detected by Western blotting using human antibody KZ51 (gift from Dennis Burton). EBOV NP truncations and mutants were detected by Western blotting using rabbit anti-2A peptide antibody ABS31 (Millipore). EBOV VP30 and VP30 mutants for the mini-replicon experiments were detected by Western blotting with a polyclonal rabbit anti-VP30 antibody (gift from Yoshihiro Kawaoka and Pete Halfmann). EBOV VP35 in the mini-replicon experiments was detected by Western blotting using a mouse anti-VP35 antibody (gift from Victor Volchkov). In co-immunoprecipitation experiments, HA-tagged VP30 proteins were detected by Western blotting using a rabbit anti-HA antibody (Rockland Immunochemicals) and FLAG-tagged VP35 proteins were detected using a rabbit anti-FLAG antibody (Cell Signaling Technology). Secondary antibodies used for Western blotting were goat anti-mouse or -rabbit or -human Fc HRP conjugates (Thermo Fisher). For immunofluorescence, HA-tagged VP30 was detected with 16B12 (BioLegend) and a goat anti-mouse Alexa 647 conjugate (Thermo Fisher).
7
Subcellular Localization of monomeric mitoTev-TALE
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COS7 cells were plated onto coverslips and transfected with each monomeric mitoTev‐TALE (1 μg total, per well of a 6‐well plate) for 24 h, as described above. In the following day, cells were stained with 200 nM MitoTracker Red CMXRos (M7512, Invitrogen, Waltham, MA) and incubated at 37°C for 1 h, protected from light. After, cells were fixed with 2% paraformaldehyde (PFA) for 15 min, at room temperature (RT). Cells were then permeabilized with a 0.2% Triton X‐100 in phosphate saline buffer (PBS) during 2 min at room temperature, followed by blocking with 3% BSA in 1× PBS during 1 h at RT. Next, the primary antibody against T2A (Millipore ABS31, 1:200 in 3% BSA/PBS) was incubated for 1 h at RT. After washing the coverslips 3× with 1× PBS, the cells were incubated with the secondary antibody Alexa Fluor 488 goat anti‐rabbit IgG (A‐11008, Invitrogen 1:200 in 3%BSA/PBS) for 1 h at RT, protected from light. The coverslips were washed with 1× PBS and placed over a DAPI‐containing mounting medium (VECTASHIELD HardSet Antifade Mounting Medium with DAPI, H‐1500 Vector Laboratories), to stain the nucleus (Hashimoto et al, 2015 ). Images were recorded in a Zeiss LSM510 confocal microscope.
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