Log In Sign up
The largest database of trusted experimental protocols

Dclk1

Manufactured by Cell Signaling Technology
Visit Supplier
Sourced in United States

DCLK1 is a protein that plays a role in cell signaling pathways. It is a member of the doublecortin-like kinase family and is involved in the regulation of microtubule dynamics.

Automatically generated - may contain errors

Lab products found in correlation

Gapdh, by Cell Signaling Technology (3 mentions) Bio plex lysis buffer, by Bio-Rad (2 mentions) Kraslsl g12d, by Jackson ImmunoResearch (2 mentions) Factor 1, by Bio-Rad (2 mentions) Factor 2, by Bio-Rad (2 mentions) Pdx1 cre mice, by Jackson ImmunoResearch (2 mentions) Tp53lsl r172h, by Jackson ImmunoResearch (2 mentions) Complete protease inhibitor cocktail, by Merck Group (2 mentions) Rm2235 microtome, by Leica (2 mentions) Permount, by Thermo Fisher Scientific (2 mentions) Target retrieval buffer, by Agilent Technologies (2 mentions) Envision hrp kit, by Agilent Technologies (2 mentions) Bx ucb slide scanner, by Olympus (2 mentions) Bcl2 associated x (bax), by Cell Signaling Technology (1 mentions) Cyclin d1, by Cell Signaling Technology (1 mentions) Stat5, by Cell Signaling Technology (1 mentions) Phospho stat5, by Cell Signaling Technology (1 mentions) Dclk1, by Merck Group (1 mentions) Phospho erk, by Cell Signaling Technology (1 mentions) Caspase 3, by Cell Signaling Technology (1 mentions)

8 protocols using dclk1

1

Pancreatic Protein Extraction and Analysis

Cited 1 time
Check if the same lab product or an alternative is used in the 5 most similar protocols
Pancreases were harvested from 8-week-old mice and flash frozen in liquid N2 immediately upon dissection. Frozen tissue was lysed in Bio-Plex Lysis Buffer (Bio-Rad, #171304011) supplemented with Factor I (Bio-Rad, #171304011), Factor II (Bio-Rad, #171304011), and cOmplete protease inhibitor cocktail (Sigma-Aldrich) according to the manufacturer’s instructions. Lysates were clarified by centrifugation and immunoblotting was performed as previously described.55 (link) The following primary antibodies were employed in this study: DCLK1 (Cell Signaling, #62257) and GAPDH (Cell Signaling, #D16H11). Immunoglobulin G horseradish peroxidase linked secondary antibodies were employed as appropriate.
Animal studies for immunohistochemistry and immunoblotting (Supplementary Fig. 8a–b) were approved and performed in accordance with the IACUC at Beth Israel Deaconess Medical Center. All experiments were adherent to institutional standards and were performed in pathogen-free animal facilities. Pdx1-Cre mice were obtained from The Jackson Laboratory (#014647); K-RasLSL-G12D (#01XJ6) and Tp53LSL-R172H (#01XM2) mice were obtained from the NCI Mouse Repository.
Ferguson F.M., Nabet B., Raghavan S., Liu Y., Leggett A.L., Kuljanin M., Kalekar R.L., Yang A., He S., Wang J., Ng R.W., Sulahian R., Li L., Poulin E.J., Huang L., Koren J., Dieguez-Martinez N., Espinosa S., Zeng Z., Corona C.R., Vasta J.D., Ohi R., Sim T., Kim N.D., Harshbarger W., Lizcano J.M., Robers M.B., Muthaswamy S., Lin C.Y., Look A.T., Haigis K.M., Mancias J.D., Wolpin B.M., Aguirre A.J., Hahn W.C., Westover K.D, & Gray N.S. (2020). Discovery of a selective inhibitor of Doublecortin Like Kinase 1. Nature chemical biology, 16(6), 635-643.
+ Open protocol
+ Expand
2

Protein Expression Analysis of Cellular Samples

Check if the same lab product or an alternative is used in the 5 most similar protocols
Cell lysates were subjected to polyacrylamide gel electrophoresis, and then blotted onto Immobilin-P polyvinylidene difluoride membranes (Millipore, Bedford, MA). Antibodies for cyclin D1 (catalog # 2978), CDK2 (catalog # 2546), phospho-Rb (catalog # 8516), Rb (catalog # 9309), PARP(catalog No # 9532), Caspase 3 (catalog # 9662), Bcl2 (catalog # 2876), BclXL (catalog # 2764), Bax (catalog No # 5023), CD44 (catalog # 3570), CD133 (catalog # 64326), phospho-STAT5 (catalog #s 4332, 9359), STAT5 (catalog # 25656), phospho-STAT3 (catalog # 9131), STAT3 (catalog # 4904), phospho-ERK (catalog # 4370) and ERK (catalog # 9102), ABCG2 (catalog #42078), DCLK1 (catalog # 62257) were purchased from Cell Signaling Technology (Danvers, MA, USA), for DCLK1 (catalog # ab37994), Oct-4 (catalog # ab189857) from Abcam Inc. (Cambridge, MA, USA), CDK4 (catalog # MA5–13498), CDK6 (catalog # MA5–13338) from Thermofisher (Waltham, MA, USA) and DCLK1 (catalog # SAB2420186) and Actin (catalog # A1978) from Sigma Aldrich and GAPDH (catalog # sc-365062) Santacruz Biotechnology Inc (Santa Cruz, CA, USA). Specific proteins were detected by chemiluminescence system.
Subramaniam D., Angulo P., Ponnurangam S., Dandawate P., Ramamoorthy P., Srinivasan P., Iwakuma T., Weir S.J., Chastain K, & Anant S. (2020). Suppressing STAT5 signaling affects osteosarcoma growth and stemness. Cell Death & Disease, 11(2), 149.
+ Open protocol
+ Expand
3

Pancreatic Protein Extraction and Analysis

Cited 1 time
Check if the same lab product or an alternative is used in the 5 most similar protocols
Pancreases were harvested from 8-week-old mice and flash frozen in liquid N2 immediately upon dissection. Frozen tissue was lysed in Bio-Plex Lysis Buffer (Bio-Rad, #171304011) supplemented with Factor I (Bio-Rad, #171304011), Factor II (Bio-Rad, #171304011), and cOmplete protease inhibitor cocktail (Sigma-Aldrich) according to the manufacturer’s instructions. Lysates were clarified by centrifugation and immunoblotting was performed as previously described.55 (link) The following primary antibodies were employed in this study: DCLK1 (Cell Signaling, #62257) and GAPDH (Cell Signaling, #D16H11). Immunoglobulin G horseradish peroxidase linked secondary antibodies were employed as appropriate.
Animal studies for immunohistochemistry and immunoblotting (Supplementary Fig. 8a–b) were approved and performed in accordance with the IACUC at Beth Israel Deaconess Medical Center. All experiments were adherent to institutional standards and were performed in pathogen-free animal facilities. Pdx1-Cre mice were obtained from The Jackson Laboratory (#014647); K-RasLSL-G12D (#01XJ6) and Tp53LSL-R172H (#01XM2) mice were obtained from the NCI Mouse Repository.
Ferguson F.M., Nabet B., Raghavan S., Liu Y., Leggett A.L., Kuljanin M., Kalekar R.L., Yang A., He S., Wang J., Ng R.W., Sulahian R., Li L., Poulin E.J., Huang L., Koren J., Dieguez-Martinez N., Espinosa S., Zeng Z., Corona C.R., Vasta J.D., Ohi R., Sim T., Kim N.D., Harshbarger W., Lizcano J.M., Robers M.B., Muthaswamy S., Lin C.Y., Look A.T., Haigis K.M., Mancias J.D., Wolpin B.M., Aguirre A.J., Hahn W.C., Westover K.D, & Gray N.S. (2020). Discovery of a selective inhibitor of Doublecortin Like Kinase 1. Nature chemical biology, 16(6), 635-643.
+ Open protocol
+ Expand
4

Protein Isolation and Western Blot Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols
Total protein from cultured A549/DDP and H1299/DDP cells was isolated in RIPA lysis buffer (Beyotime, Shanghai, China). According to Bradford protein assay reagent (Bio-Rad), equal amounts of protein (20 μg) from each sample were loaded for the standard procedures of Western blot assay. β-actin was used to normalize the DCLK1 protein level. The primary antibodies including DCLK1 (#62,257, 1:1000) and β-actin (#4967, 1:1000) were purchased from Cell Signaling Technology (CST; Danvers, MA, USA), and Ki-67 (#ab197547, 1:500), cleaved caspase-3 (#ab49822, 1:500) were from abcam (Cambridge, UK).
Zhan Y., Abuduwaili K., Wang X., Shen Y., Nuerlan S, & Liu C. (2020). Knockdown of Long Non-Coding RNA HOTAIR Suppresses Cisplatin Resistance, Cell Proliferation, Migration and Invasion of DDP-Resistant NSCLC Cells by Targeting miR-149-5p/Doublecortin-Like Kinase 1 Axis. Cancer Management and Research, 12, 7725-7737.
+ Open protocol
+ Expand
5

Immunohistochemical Analysis of DCLK1 in Murine Pancreas

Check if the same lab product or an alternative is used in the 5 most similar protocols
Pancreases were harvested from mice at indicated ages and fixed in 10% formalin overnight at room temperature. Fixed tissue was processed into paraffin blocks and cut into 5 μM sections using a Leica RM2235 microtome. Tissue sections were deparaffinized in a xylene and ethanol series and then washed with water. Antigen retrieval was performed by incubating slides in Target Retrieval Buffer (DAKO, #S1699) in a pressure cooker. Immunohistochemical analysis was performed using the EnVision+ HRP Kit (DAKO, #401111) and DCLK1 (Cell Signaling, #62257) according the manufacturer’s instructions. Slides were counterstained with hematoxylin, dehydrated through an ethanol and xylene series, and coverslips were mounted using Permount (Fisher, #SP15–100). Images were acquired using an Olympus BX-UCB slide scanner.
Ferguson F.M., Nabet B., Raghavan S., Liu Y., Leggett A.L., Kuljanin M., Kalekar R.L., Yang A., He S., Wang J., Ng R.W., Sulahian R., Li L., Poulin E.J., Huang L., Koren J., Dieguez-Martinez N., Espinosa S., Zeng Z., Corona C.R., Vasta J.D., Ohi R., Sim T., Kim N.D., Harshbarger W., Lizcano J.M., Robers M.B., Muthaswamy S., Lin C.Y., Look A.T., Haigis K.M., Mancias J.D., Wolpin B.M., Aguirre A.J., Hahn W.C., Westover K.D, & Gray N.S. (2020). Discovery of a selective inhibitor of Doublecortin Like Kinase 1. Nature chemical biology, 16(6), 635-643.
+ Open protocol
+ Expand
6

Immunohistochemical Analysis of DCLK1 in Murine Pancreas

Check if the same lab product or an alternative is used in the 5 most similar protocols
Pancreases were harvested from mice at indicated ages and fixed in 10% formalin overnight at room temperature. Fixed tissue was processed into paraffin blocks and cut into 5 μM sections using a Leica RM2235 microtome. Tissue sections were deparaffinized in a xylene and ethanol series and then washed with water. Antigen retrieval was performed by incubating slides in Target Retrieval Buffer (DAKO, #S1699) in a pressure cooker. Immunohistochemical analysis was performed using the EnVision+ HRP Kit (DAKO, #401111) and DCLK1 (Cell Signaling, #62257) according the manufacturer’s instructions. Slides were counterstained with hematoxylin, dehydrated through an ethanol and xylene series, and coverslips were mounted using Permount (Fisher, #SP15–100). Images were acquired using an Olympus BX-UCB slide scanner.
Ferguson F.M., Nabet B., Raghavan S., Liu Y., Leggett A.L., Kuljanin M., Kalekar R.L., Yang A., He S., Wang J., Ng R.W., Sulahian R., Li L., Poulin E.J., Huang L., Koren J., Dieguez-Martinez N., Espinosa S., Zeng Z., Corona C.R., Vasta J.D., Ohi R., Sim T., Kim N.D., Harshbarger W., Lizcano J.M., Robers M.B., Muthaswamy S., Lin C.Y., Look A.T., Haigis K.M., Mancias J.D., Wolpin B.M., Aguirre A.J., Hahn W.C., Westover K.D, & Gray N.S. (2020). Discovery of a selective inhibitor of Doublecortin Like Kinase 1. Nature chemical biology, 16(6), 635-643.
+ Open protocol
+ Expand
7

Western Blot Analysis of Intestinal Proteins

Check if the same lab product or an alternative is used in the 5 most similar protocols
Isolated enterocytes were lysed with sodium dodecyl sulfate sample buffer (50 mM Tris-HCl pH 6.8, 2% sodium dodecyl sulfate, 10% glycerol, and 5% mercaptoethanol). The lysate was then heated for 5 min at 100 °C, and the protein concentration was determined using the RC-DC kit (Bio-Rad, Hercules CA). Proteins were loaded in equal amounts for Western blotting. Antibodies used in this study were Raptor (#2280), p-S6K (#9234), pS6 (S240/244) (#5364), DCLK1 (#62257), Stat 6 (#5397), GAPDH (#5174) (all from Cell Signaling Technology, Danvers MA) and p-Stat 6 (sc-11762, Santa Cruz Biotechnology, Dallas Texas). The proteins were detected using Bio-Rad ChemiDocTM XRS + system with image Lab TM software (Bio-Rad, Hercules CA).
Aladegbami B., Barron L., Bao J., Colasanti J., Erwin C.R., Warner B.W, & Guo J. (2017). Epithelial cell specific Raptor is required for initiation of type 2 mucosal immunity in small intestine. Scientific Reports, 7, 5580.
+ Open protocol
+ Expand
8

Automated Immunohistochemistry Protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols
IHC was performed using an automated immunostaining system (BONDMAX; Leica Biosystems, Melbourne, Australia). Briefly, to begin with, the samples were rinsed with xylene for deparaffinization. The slides were rehydrated using increasing concentrations of alcohol and washed with phosphate-buffered saline. Antigens were retrieved and incubated with a bond peroxidase-blocking reagent, followed by primary antibodies against AR (1:200 dilution, Cell Signaling), CD8 (1:500, Cell Signaling), FOXC1 (1:200, Abcam), and DCLK1 (1:100, Cell Signaling Technology). A Bond Polymer Refining Detection Kit (Leica) was used for detection, and 3,3′-diaminobenzidine was used for immunodetection, followed by Mayer’s hematoxylin for counterstaining. We used prostate tissue and tonsil as positive controls for AR and CD8 expression, respectively. TNBC tissues which were tested strongly positive were used as positive control for DCLK1 and FOXC1 expression. The positive control tissues were put simultaneously on all of the examined slides.
Leeha M., Kanokwiroon K., Laohawiriyakamol S, & Thongsuksai P. (2023). Immunohistochemistry-based molecular subtyping of triple-negative breast cancer and its prognostic significance. Pathology and Oncology Research, 29, 1611162.
+ 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!