The following antibodies were used for mouse tissues immunohistochemistry and immunofluorescence: Mouse anti-Human TDP-43 (Novus 2E2–D3); Rabbit anti-Mouse TDP-43 N-Term (Proteintech 10782–2); Rabbit anti-Mouse TDP-43 C-Term (Proteintech 12892–1);Rabbit anti-Synaptophysin (Life Technologies Z66, 1:500); Mouse anti-Pan-Axonal neruofilament (Covance, SMI-312R, 1:1000). The following antibodies were used for cortex and spinal cord immunoblot: Mouse anti-Human TDP-43 (Novus 2E2–D3); Rabbit anti-Mouse TDP-43 N-Term (Proteintech 10782–2); Rabbit anti-Mouse TDP-43 C-Term (Proteintech 12892–1); Mouse anti-GAPDH (Sigma, GAPDH-71.1).The following antibodies were used for i3Neuron stable cell lines immunoblot: Rabbit anti-TDP-43 (Proteintech 10782–2-AP 1:1000); Mouse anti-FLAG (Sigma F1804, 1:5000); Rabbit anti-GAPDH (CST 2118, 1:1000); Mouse anti-Tubulin (Thermofisher 14–4502–82, 1:1000). The following antibodies were used for QBI-293 stable cell lines immunoblot: Mouse anti-TDP-43 (abcam ab104223, 1:2000); Rabbit anti-GFP (Cell signalling mAb 2956, 1:1000); Mouse anti-Beta Actin (Proteintech 66009–1, 1:2000). The following antibody was used for HEK-293 cells immunoblot: Rabbit anti-Mouse TDP-43 N-Term (Proteintech 10782–2, 1:2000).
Mouse anti tubulin
Manufactured by Thermo Fisher Scientific
Sourced in United States
Mouse anti-tubulin is a primary antibody that recognizes the tubulin protein, a key component of the cytoskeleton. It can be used to detect and visualize tubulin in various cell and tissue samples.
Automatically generated - may contain errors
Lab products found in correlation
Trans blot turbo transfer system, by Bio-Rad (3 mentions)
Nitrocellulose membrane, by Thermo Fisher Scientific (2 mentions)
Lsm 700 confocal microscope, by Zeiss (2 mentions)
Rat anti klp 18, by Thermo Fisher Scientific (2 mentions)
Anti rat hrp, by Thermo Fisher Scientific (2 mentions)
Clarity western ecl substrate, by Bio-Rad (2 mentions)
Mouse anti gapdh, by Merck Group (1 mentions)
Mouse anti human tdp 43, by Proteintech (1 mentions)
Rabbit anti mouse tdp 43 n term, by Proteintech (1 mentions)
Rabbit anti mouse tdp 43 c term, by Proteintech (1 mentions)
Rabbit anti synaptophysin, by Thermo Fisher Scientific (1 mentions)
Mouse anti human tdp 43, by Novus Biologicals (1 mentions)
Rabbit anti tdp 43, by Proteintech (1 mentions)
Mouse anti tdp 43, by Abcam (1 mentions)
Rabbit anti gfp, by Cell Signaling Technology (1 mentions)
Mouse anti beta actin, by Proteintech (1 mentions)
Mouse anti flag, by Merck Group (1 mentions)
Smi 312r, by Fortrea (1 mentions)
Superblock buffer, by Thermo Fisher Scientific (1 mentions)
Criterion tris hcl gel, by Bio-Rad (1 mentions)
10 protocols using mouse anti tubulin
1
Antibodies for TDP-43 Immunohistochemistry and Immunoblotting
2
Western Blot Analysis of Protein Lysates
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
An aliquot of the total lysates was used for western blotting (WB). 0.1% SDS was added to the lysates, followed by sonication (3 cycles, 10 s each). Samples were run on 4–20% Criterion Tris-HCl gels (Bio-Rad Laboratories) and electrophoretically transferred to a nitrocellulose membrane (Thermo Fisher Scientific). Residual protein-binding sites were blocked by incubation with 5% non-fat milk in 1XTBST (1X TBS plus 0.1% Tween 20) 1 h at RT, followed by an overnight (O/N) incubation at 4 °C with primary antibodies diluted in 20% SuperBlock buffer (Thermo Fisher Scientific) in 1XTBST. Mouse anti-tubulin (Thermo Fisher Scientific) were diluted 1:5000; anti-Myc-tag 9B11 (Cell Signaling, Danvers, MA) was diluted 1:1000. Appropriate isotypes secondary antibodies HRP-conjugated were diluted 1:2000 or 1:10000 in 5% non-fat milk 1XTBST, and added for 1 h at RT. Every step was followed by 3 or 4 washes in 1X TBST. Detection was performed using Pierce ECL Western Blotting Substrate (Thermo Fisher Scientific) or SuperSignal West Dura extended duration substrate (Thermo Fisher Scientific) and exposed to x-ray films.
3
Larval CNS Protein Extraction and Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Larval central nervous system (CNS) protein extractions were performed as follows: 10 CNSs were dissected from third instar larvae. The resulting tissues were sheared manually in 5 µl of 2% SDS aqueous solution using a micropistil secured into a 1.5-ml cup. Subsequently, 0.5 µl of a 10% Triton X-100 aqueous solution and 5 µl of 2× Laemmli sample buffer were added. Samples were then heated at 95°C for 10 min. Centrifugation was performed for 2 min at 16,000 g, in order to pellet debris. For larval CNS, 10 µl (equivalent to 10 larval CNS) was subjected to denaturing SDS-PAGE using a 6% Tris-HCl gel.
Proteins of the PAGE gel were then transferred onto a nitrocellulose membrane blocked with 5% skim milk in 1× PBS supplemented with 0.1% Tween-20 and probed with rabbit anti-Rab2 (1:5,000; Santa Cruz Biotechnology; sc-285667) and mouse anti-tubulin (1:100,000; Thermo Fisher Scientific; 62204) diluted in 5% milk in 1× PBS, supplemented with 0.1% Tween-20. After washing, secondary anti-rabbit or anti-mouse IgG HRP-conjugated antibodies (Dianova) were used for detection (Dianova) in conjunction with an enhanced chemoluminescence (GE Healthcare ECL Prime; RPN 2232) detection system with Hyperfilm ECL (GE Healthcare).
Proteins of the PAGE gel were then transferred onto a nitrocellulose membrane blocked with 5% skim milk in 1× PBS supplemented with 0.1% Tween-20 and probed with rabbit anti-Rab2 (1:5,000; Santa Cruz Biotechnology; sc-285667) and mouse anti-tubulin (1:100,000; Thermo Fisher Scientific; 62204) diluted in 5% milk in 1× PBS, supplemented with 0.1% Tween-20. After washing, secondary anti-rabbit or anti-mouse IgG HRP-conjugated antibodies (Dianova) were used for detection (Dianova) in conjunction with an enhanced chemoluminescence (GE Healthcare ECL Prime; RPN 2232) detection system with Hyperfilm ECL (GE Healthcare).
4
Immunofluorescence Staining of Organelle Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
COS-7 cells or U2OS cells were fixed with 4% paraformaldehyde (PFA) in PBS for 25 min, permeabilized with 0.1% Triton X-100/PBS for 10 min, and blocked with 3% BSA for 1 h at room temperature. Fixed cells were then incubated with primary antibodies for 1 h at room temperature or overnight at 4 °C, including rabbit anti-calreticulin (Abcam; 1:800), mouse anti-Tubulin (Thermo; 1:200), rabbit anti-Tubulin (Abcam; 1:1000), mouse anti-HA (Sigma; 1:500), rabbit anti-HA (Abcam; 1:1000), mouse anti-GM130 (BD; 1:500) and mouse anti-TOM20 (BD; 1:500), followed by incubation with various fluorophore-conjugated secondary antibodies (Alexa Fluor 488-conjugated anti-rabbit or mouse, Alexa Fluor 568-conjugated anti-mouse or rabbit, Invitrogen) for 1 h at room temperature. All images were captured on Leica TCS SP5 or Zeiss LSM700 confocal microscope with a 63× objective. Brightness and contrast were adjusted across the entire image using Adobe Photoshop.
5
Western Blot Analysis of Sima and Msi Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Protein extracts from S2R+ cells or embryos (stage E14 to stage E17) were prepared in RIPA buffer (50 mM Tris–HCl pH 7.5, 150 mM NaCl, 0.1% (v/v) SDS, 0.5% (w/v) sodium deoxycholate and 1% Triton X-100) with the addition of proteinase inhibitory cocktail (Invitrogen) and kept at 4°C. 25–50 μg of total extracts were loaded on a 6–11% polyacrylamide gel, subjected to electrophoresis and then blotted onto nitrocellulose membrane (Bio-Rad). Thereafter, membranes were blocked for 1 h at room temperature with 5% nonfat milk or BSA in TBS with 0.1% Tween 20 (TBS-T) and incubated overnight with rabbit anti-Sima (29 (link)); rat anti-dMsi 3A5 (26 (link)) or mouse anti-tubulin (Invitrogen) in 5% nonfat milk in TBS-T. The secondary antibodies used were HRP conjugated (1/5000, Jackson ImmunoResearch). Immunoblots were developed with the ECL prime detection reagent (Amersham).
6
Tracking Protein Expression in Transfected BHK-21 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
BHK-21 cells were transfected with RNA transcripts from pGFP-PAC or pLL-GFP-PAC, cell extracts prepared at 1, 2, 4, 6, 8, 10 and 24 h post-transfection using RIPA lysis buffer (150 mM NaCl, 50 mM Tris–HCl [pH 7.4], 1% NP-40, 0.5% deoxycholic acid, 0.1% SDS). Samples were resolved by 10% SDS-PAGE, transferred to a nitrocellulose membrane (Life Technologies) and blocked for 1 h in 5% PBS-T (PBS, 0.1% Tween 20, supplemented with 5% non-fat milk). The membrane was probed with rabbit anti-eIF4G (1:1000: kind gift of Lisa Roberts) or mouse anti-tubulin (Invitrogen, 1:2000) overnight at 4 °C. The membrane was then washed 3× in PBS-T, then probed with peroxidase conjugated anti-rabbit (Invitrogen, 1:2000) or anti-mouse (Invitrogen, 1:2000) antibodies in 5% PBS-T for 1 h at room temperature. Membranes were washed 3× in PBS-T and antibody binding detected using an EZ-ECL HRP chemiluminescence detection kit Biological Industries, Kibbutz Beit-Haemek, Israel).
7
Immunolocalization of CoQ6 and PDSS2 in Oocytes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Ovarian sections fixed in 10% neutral buffered formalin were embedded in paraffin, sectioned, and rehydrated. Antigen retrieval with citrate buffer was used prior to exposure to antibody PDSS2 (1:40) followed by ABC Vectastain Kit (Dako, Glostrup, Denmark). Sections from Pdssfl/fl ZP3-Cre+ females were used as a negative control. Isolated oocytes at the GV stage were fixed in PHEM fixative, and indirect immunocytochemistry of the oocytes was performed as previously described (Fernandes et al., 2012 (link)). Primary antibodies included mouse antitubulin (1:500; Invitrogen) and rabbit anti-CoQ6 or PDSS2 (1:100; 1:200, respectively; Proteintech Group Inc, Chicago, IL, USA), followed by incubation with donkey anti-mouse Alexa 488 or donkey anti-rabbit Alexa 594 (Invitrogen). DNA was counterstained with 4′,6-diamidino-2-phenylindole (DAPI) and imaged on confocal microscope. Values obtained from samples without primary antibody were subtracted from the experimental signals, and the data were shown as the sum of intensity expressed as random fluorescence units (RFUs).
8
SARS-CoV-2 Spike Protein Detection
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were lysed in lysis buffer (10 mM iodoacetamide [Sigma-Aldrich], Complete protease inhibitor tablets [Roche], 300 mM sodium chloride, 50 mM Tris-HCl [pH 7.5], and 0.5% Triton X-100 [Sigma-Aldrich]). Virus was pelleted through 20% sucrose and resuspended in lysis buffer. Cell and virus lysates were heated to 95 °C for 5 min. Samples were analyzed on 10% 1.5 mm Tris-glycine gels, followed by transfer to PVDF membrane (BioRad) using a BioRad Trans-Blot Turbo Transfer system according to manufacturer's instructions. Blots were probed with the following antibodies: rabbit anti-SARS-CoV-2 S (Sino Biological; #40589), rabbit anti-SARS-CoV-2 S2 (Sino Biological; #40590), human anti-HIV immune globulin (HIV-Ig; NIH AIDS Reagent Program), and mouse anti-tubulin (Invitrogen; #62204). Secondary antibodies conjugated with horseradish peroxidase were used for chemiluminescent detection for SARS-CoV-2 S and other proteins were detected using near-IR fluorescently labeled secondary antibodies (Azure Biosystems 650 and 800; AC2165 and AC2135). Protein bands were visualized using a Sapphire Biomolecular Imager (Azure Biosystems) and analyzed with AzureSpot (Azure Biosystems).
9
Whole-worm Western Blots for KLP-18
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For klp-18(or447ts) whole-worm Western blots, control (OD868) or klp-18(or447ts) (SMW36) plates were shifted to 26°C for 1 h. Fifty to one hundred worms were picked from a shifted plate to a prewarmed unseeded plate for 5 min to avoid transfer of bacteria and then washed off the plate with room temperature M9. Worms were pelleted by spinning at 800 × g for 1 min, supernatant was removed, and an equal volume of 2× SDS Laemmli sample buffer was added. Gel samples were boiled for 10 min at 95°C, briefly vortexed, and then boiled for an additional 10 min at 95°C. A volume of sample corresponding to 50 worms was loaded onto the gel. Antibodies used: rat anti–KLP-18 1:1000, anti–rat-HRP 1:5000 (Invitrogen), mouse anti-tubulin 1:5000, and anti–mouse-HRP 1:5000 (Invitrogen).
Samples were run on 8–12% SDS–PAGE gel and transferred to a nitrocellulose membrane using a Trans-Blot Turbo Transfer System (BioRad). Membrane was blocked in 5% milk in TBST (TBS [20 mM Tris Base pH 7.6, 150 mM NaCl] + 0.1% Tween) blocking solution, incubated with primary antibody in blocking solution at room temperature for 1 h or at 4°C overnight, washed in TBST, incubated in secondary antibody at room temperature for 1 h, washed in TBST, incubated with Clarity Western ECL substrate (BioRad) for 2 min, and then imaged.
Samples were run on 8–12% SDS–PAGE gel and transferred to a nitrocellulose membrane using a Trans-Blot Turbo Transfer System (BioRad). Membrane was blocked in 5% milk in TBST (TBS [20 mM Tris Base pH 7.6, 150 mM NaCl] + 0.1% Tween) blocking solution, incubated with primary antibody in blocking solution at room temperature for 1 h or at 4°C overnight, washed in TBST, incubated in secondary antibody at room temperature for 1 h, washed in TBST, incubated with Clarity Western ECL substrate (BioRad) for 2 min, and then imaged.
10
Western Blot Analysis of KLP-18 in C. elegans
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For klp-18(or447ts) whole worm Western blots, control (OD868) or klp-18(or447ts) (SMW36)
plates were shifted to 26°C for 1 hour. 50-100 worms were picked from shifted plate to prewarmed unseeded plate for 5min to avoid transfer of bacteria, then washed off plate with room temperature M9. Worms were pelleted by spinning at 800xg for 1min, supernatant was removed, and an equal volume of 2X SDS Laemmli Sample Buffer was added. Gel samples were boiled for 10min at 95°C, briefly vortexed, then boiled for an additional 10min at 95°C.
Volume of sample corresponding to 50 worms was loaded onto gel. Antibodies used: rat anti-KLP-18 1:1000, anti-rat-HRP 1:5000 (Invitrogen), mouse anti-tubulin 1:5000, anti-mouse-HRP 1:5000 (Invitrogen).
Samples were run on 8-12% SDS-PAGE gel and transferred to a nitrocellulose membrane using a Trans-Blot Turbo Transfer System (BioRad). Membrane was blocked in 5% milk in TBS + 0.1% Tween blocking solution, incubated with primary antibody in blocking solution at room temperature for 1 hour or at 4°C overnight, washed in TBST, incubated in secondary antibody at room temperature for 1 hour, washed in TBST, incubated with Clarity Western ECL substrate (BioRad) for 2 minutes, then imaged. experiment: Shift to pellet (%) +/-sd for C-stalk (-MT) = 6.0 +/-6.7, C-stalk (+ MT) = 50.8% +/-7.0%, C-stalk (+ sMT) = 26.7% +/-7.0%.
plates were shifted to 26°C for 1 hour. 50-100 worms were picked from shifted plate to prewarmed unseeded plate for 5min to avoid transfer of bacteria, then washed off plate with room temperature M9. Worms were pelleted by spinning at 800xg for 1min, supernatant was removed, and an equal volume of 2X SDS Laemmli Sample Buffer was added. Gel samples were boiled for 10min at 95°C, briefly vortexed, then boiled for an additional 10min at 95°C.
Volume of sample corresponding to 50 worms was loaded onto gel. Antibodies used: rat anti-KLP-18 1:1000, anti-rat-HRP 1:5000 (Invitrogen), mouse anti-tubulin 1:5000, anti-mouse-HRP 1:5000 (Invitrogen).
Samples were run on 8-12% SDS-PAGE gel and transferred to a nitrocellulose membrane using a Trans-Blot Turbo Transfer System (BioRad). Membrane was blocked in 5% milk in TBS + 0.1% Tween blocking solution, incubated with primary antibody in blocking solution at room temperature for 1 hour or at 4°C overnight, washed in TBST, incubated in secondary antibody at room temperature for 1 hour, washed in TBST, incubated with Clarity Western ECL substrate (BioRad) for 2 minutes, then imaged. experiment: Shift to pellet (%) +/-sd for C-stalk (-MT) = 6.0 +/-6.7, C-stalk (+ MT) = 50.8% +/-7.0%, C-stalk (+ sMT) = 26.7% +/-7.0%.
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!