Cyclin d1

Manufactured by Affinity Biosciences

Sourced in United States, China

Cyclin D1 is a laboratory equipment product that plays a core role in cell cycle regulation. It is a regulatory protein involved in the progression of cells through the G1 phase of the cell cycle.

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Lab products found in correlation

14 protocols using cyclin d1

Molecular Mechanisms of LPS-Induced Inflammation

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Reagents: ampicillin (A8180-2, Solarbio, China); colistin (R002831, Rhawn, China); neomycin and vancomycin (N814740 and V820413, Macklin, China); LPS (L8880, Solarbio, China); BAY-11-7082 (S1523, Beyotime, China); Stattic (A2224, APExBIO, USA); Docetaxel (A4394, APExBIO, USA); DMSO (D8370, Solarbio, China); PEG300 (IP9020, Solarbio, China); Tween-80 (T8360, Solarbio, China); DAPI (C0065, Solarbio, China); DAB (ZLI-9018, ZSGB-BIO, China).
Antibodies: Ki-67 (ab15580, abcam, UK); LPS (MAB526Ge22, Cloud-Clone, China); p65 (AF1234, Beyotime, China); p-p65 (Ser536) (AF2006, Affinity, USA); p-STAT3 (Tyr705) (AF3293, Affinity, USA); STAT3 (AF1492, Beyotime, China); Antibody-IL6 (AF-406-SP, R&D, USA); C-myc (AF0358, Affinity, USA); Cyclin D1 (AF0931, Affinity, USA); Bcl-2 (AF6285, Beyotime, China); Survivin (AF6017, Affinity, USA); β-actin (AF5003, Beyotime, China); β-tubulin (AF1216, Beyotime, China). Second antibodies: HRP-conjugated goat anti-rabbit IgG (cw0103s, CWBIO, China); HRP-conjugated goat anti-mouse IgG (cw0102s, CWBIO, China); fluorescent Cy3-conjugated goat anti-rabbit IgG (BA1032, BOSTER, China).

Zhong W., Wu K., Long Z., Zhou X., Zhong C., Wang S., Lai H., Guo Y., Lv D., Lu J, & Mao X. (2022). Gut dysbiosis promotes prostate cancer progression and docetaxel resistance via activating NF-κB-IL6-STAT3 axis. Microbiome, 10, 94.

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Protein Expression Profile Analysis

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Total proteins were extracted using radioimmunoprecipitation assay (RIPA) buffer supplemented with protease inhibitor Cocktail (Roche, Switzerland). Total cell lysates were separated by 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to polyvinylidene fluoride (PVDF) membranes (Roche, Switzerland). Membranes were blocked with 5% bovine serum albumin (BSA) (Sigma-Aldrich, USA) at room temperature for 1h, and then incubated overnight at 4 °C with primary antibodies included Cyclin D1 (1/2000) (Affinity, USA), C-myc (1/1000) (Affinity, USA), MMP2 (1/1000) (Boster, China), E-cadherin (1/1000) (BD Biosciences) and β-actin (1/500) (Boster, China). Then, the membranes were incubated with secondary antibody after three times wash and visualized with enhanced chemiluminescence (ECL) assay kit (Millipore, USA).

Zhang Q., Wang C., Miao S., Li C., Chen Z, & Li F. (2017). Enhancing E-cadherin expression via promoter-targeted miR-373 suppresses bladder cancer cells growth and metastasis. Oncotarget, 8(55), 93969-93983.

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Western Blot Analysis of Cell Signaling Proteins

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The total protein was extracted with a RIPA lysis buffer, and its concentration was determined with a BCA kit (Solarbio). The polyacrylamide gel was prepared in advance, and its concentration was determined by the size of the protein to be detected. The protein sample was loaded into a polyacrylamide gel, followed by electrophoresis for 2–3 h. Afterward, the protein was transferred onto a polyvinylidene fluoride membrane (Millipore, Billerica, MA, USA), and skim milk was used to block the heterogenetic antigens. Subsequently, the membrane loaded with protein was incubated with primary antibody at 4°C in the dark overnight. After rinsing with TBST buffer, the membrane was incubated with a secondary antibody labeled with HRP, and reacted with ECL reagent (Solarbio) for several minutes, followed by signal exposure in the dark. The antibody information was shown in the following: RASIP1 (1:1000; Affinity, Changzhou, Jiangsu, China), cyclin E1 (1:1000; Affinity), cyclin B1 (1:1000; Affinity), cyclin D1 (1:1000; Affinity), p27 (1:1000; Affinity), matrix metalloproteinase 2 (MMP2) (1:2000; Novus Biologicals, Littleton, CO, USA), MMP9 (1:1000; Affinity), cleaved caspase-3 (1:1000; Affinity), Bax (1:1000; Affinity), Bad (1:1000; Affinity), Bcl-xl (1:1000; Affinity), FOXO3 (1:500; Affinity), GAPDH (1:10000; Affinity).

Xing X., Wang X., Liu M., Guo Q, & Wang H. (2023). Ras interacting protein 1 facilitated proliferation and invasion of diffuse large B-cell lymphoma cells. Cancer Biology & Therapy, 24(1), 2193114.

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Western Blot Analysis of Protein Expression

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With respect to the detection of protein expression, Western blot was carried out as previously described [24] (link). After 48 h of transfection, cells were harvested for analysis of protein expression. In brief, a total of 20 μg protein extract from tissues and cells was loaded onto polyacrylamide gels and transferred onto nitrocellulose membranes prior to blocking aspecific signals using defatted dry milk. Then, the membranes were incubated with the antibodies of proliferating cell nuclear antigen (PCNA; 1:1000; Thermo Fisher), B-cell lymphoma 2 (Bcl-2; 1:500; Thermo Fisher), Bcl-2-like protein 4 (Bax; 1:6000; Thermo Fisher), TRIM66 (1:1000; Thermo Fisher), β-catenin (1:2000; Affinity, Nanjing, China), c-myc (1:1000; Thermo Fisher), cyclinD1 (1:2000; Affinity), MMP2 (1:3000; Abcam, Cambridge, UK) and MMP9 (1:5000; Abcam) and GAPDH (1:400; Thermo Fisher). The blots were probed with secondary antibodies (1:4000; Thermo Fisher). At last, protein visualization was achieved using eyoECL Plus (Beyotime, Shanghai, China).

Wang W., Wang J., Li Y, & Zhao Y. (2022). Circ_0051079 silencing inhibits the malignant phenotypes of osteosarcoma cells by the TRIM66/Wnt/β-catenin pathway in a miR-625-5p-dependent manner. Journal of Bone Oncology, 35, 100436.

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Western Blot Analysis of Protein Expression

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Total tissues and cellular proteins were extracted and separated by 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and then transferred onto polyvinylidene fluoride membranes (Millipore, USA). The membranes incubated overnight at 4 °C with primary antibodies. The antibodies against CDK2, Cyclin D1, β-catenin, C-myc, N-cadherin, Snail were purchased from Affinity (USA), p21 antibody was from Cell Signaling Technology (USA), E-cadherin antibody was from BD Biosciences (USA), antibodies against MMP9, Vimentin, β-actin and GAPDH were from Boster (China).

Zhang Q., Miao S., Han X., Li C., Zhang M., Cui K., Xiong T., Chen Z., Wang C, & Xu H. (2018). MicroRNA-3619-5p suppresses bladder carcinoma progression by directly targeting β-catenin and CDK2 and activating p21. Cell Death & Disease, 9(10), 960.

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Characterization of ALV-J Infection in Cell Lines

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The 293 T, LMH and DF-1 cell lines were kept in our laboratory. The 293 T and DF-1 cell lines were cultured in dulbecco’s modified eagle medium (DMEM), and LMH cells were cultured in DMEM/F12 (Gibco, Thermo Fisher Scientific, Inc.) with 10% fetal bovine serum (FBS) and maintained at 37 °C with 5% CO₂. The chronic transformed ALV-J Hc1 strain was also kept in our laboratory.
The primary antibodies of β-catenin and c-Myc were the products of Santa Cruz Biotechnology (Santa Cruz, CA, USA). The primary antibodies of TCF4 and Cyclin D1 were the products of Affinity Biosciences, Ltd. (USA). An GAPDH primary antibody was the product of Abcam, Inc. (UK). An eGFP-Tag primary antibody was the product of Invitrogen, Thermo Fisher Scientific, Inc. (California, USA). A FLAG-Tag primary antibody was the product of Sigma-Aldrich, St. Louis, Missouri (USA). The mouse monoclonal antibodies G2.3 and JE9 against the gp85 protein of ALV-J was kindly provided by the Avian Disease and Oncology Laboratory (USA) and Professor Aijian Qin of Yangzhou University, respectively. The single factor serum of chTERT protein was prepared and preserved by our laboratory in the early stage.

Xiang Y., Liang C., Li Q., Chen Q., Zhou Y., Zheng X., Zhou D., Wang Z., Wang G, & Cao W. (2022). Chicken telomerase reverse transcriptase promotes the tumorigenicity of avian leukosis virus subgroup J by regulating the Wnt/β-catenin signaling pathway. Veterinary Research, 53, 100.

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Neoprzewaquinone A Cytotoxicity Evaluation

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Neoprzewaquinone A (purity ≥98%) was obtained from Herbpurify (Chengdu, China). SGI-1776 and Netarsudil (purity ≥98%) were purchased from Selleck (Chengdu, China). RPMI 1640, DMEM, DMEM/F12 media and Fetal Bovine Serum (FBS) were acquired from Gibco (NewYork, NY, USA). Paraformaldehyde (PFA; 4%) was purchased from Biosharp (Hefei, China). The BCA protein assay kit, cell cycle analysis kit, annexin V-FITC/PI apoptosis detection kit and Hoechst 33258 were obtained from Beyotime (Shanghai, China). The trypsin, crystal violet (1%), and Mondansylcadaverine (MDC) assay kit were purchased from Solarbio (Beijing, China). The primary antibodies against ROCK1, ROCK2, mTOR/p-mTOR, MYPT1/p-MYPT1, PIM1, BAD/p-BAD, STAT3/p-STAT3, E-cadherin, Vimentin, cyclin B1, cyclin D1, Beclin1, ATG5, LC3B, GAPDH, β-actin, and HRP-conjugated secondary antibodies were purchased from Affinity (Jiangsu, China). The electrochemiluminescence (ECL) western blot detection kit was purchased from Abbkine (Redlands, CA, USA).

Zhao G., Ren Y., Yan J., Zhang T., Lu P., Lei J., Rao H., Kang X., Cao Z., Peng F., Peng C., Rao C, & Li Y. (2023). Neoprzewaquinone A Inhibits Breast Cancer Cell Migration and Promotes Smooth Muscle Relaxation by Targeting PIM1 to Block ROCK2/STAT3 Pathway. International Journal of Molecular Sciences, 24(6), 5464.

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Byakangelicin and Silibinin Modulate Liver Fibrosis

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Byakangelicin (purity ≥ 98.5%) was obtained from Push Bio‐technology Co., Ltd. Silibinin (purity > 97%) was obtained from Meilunbio Co., Ltd. Olive oil and carbon tetrachloride were obtained from Shanghai Macklin Biochemical Co., Ltd. byakangelicin was dissolved in dimethylsulphoxide (DMSO) from Sigma‐Aldrich Corporation for cell experiment. Recombinant human PDGF‐BB was obtained from R&D system. Recombinant Human TGF‐β1 was obtained from PeproTtech. Primary antibodies including anti‐α‐smooth muscle actin (α‐SMA), anti‐collagen Ⅰ (COL‐1), anti‐GAPDH and anti‐p‐Stat3, Stat3, β‐tubulin, cyclin D1, P53, p‐ASK‐1, ASK‐1, IL‐1β, NF‐κB and Nrf‐2 were acquired from Affinity Biosciences; anti‐p‐Smad3, Smad3, p‐PDGFR, PDGFR, p‐ERK, ERK, p‐AKT, AKT, PARP, cleaved caspase‐3 and caspase‐3 were received from (Cell Signaling Technology); anti‐p‐JNK, JNK from Signalway Antibody LLC; and anti‐4‐HNE from Abcam. Reagents and antibodies are prepared according to the instructions.

Li X., Shao S., Li H., Bi Z., Zhang S., Wei Y., Bai J., Zhang R., Ma X., Ma B., Zhang L., Xie C., Ning W., Zhou H, & Yang C. (2020). Byakangelicin protects against carbon tetrachloride–induced liver injury and fibrosis in mice. Journal of Cellular and Molecular Medicine, 24(15), 8623-8635.

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FGF21 Regulation of Mitochondrial Function

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FGF21 was purchased from PeproTech. The miR‐130 agomir was obtained from RiboBio. The primary antibodies used in this study included PPARγ (Abcam, #ab178860), α‐SMA (Abcam, #ab5694), bcl‐2 (Abcam, #ab59348), proliferating cell nuclear antigen (PCNA; Cell Signalling Technology, #13110), β‐actin (Cell Signalling Technology, #4970), Bax (Cell Signalling Technology, #14796), caspase‐3 (Cell Signalling Technology, #9665), cleaved caspase‐3 (Cell Signalling Technology, #9665), apoptosis‐inducing factor (AIF; Cell Signalling Technology, #5318), cytochrome c (Cyt C; Cell Signalling Technology, #11940), cyclin‐dependent kinase 1 (CDK1; Affinity Biosciences, DF6024), cyclin D1 (Affinity Biosciences, AF0931) and COX IV (Affinity Biosciences, AF5468). The PPARγ agonist pioglitazone (Pio) was obtained from Selleck (Houston, TX, USA). Donkey anti‐mouse IgG H&L (Alexa Fluor 488; lot no. ab150105) antibodies were purchased from Abcam. Horseradish peroxidase (HRP)‐conjugated goat anti‐rabbit IgG antibody (lot no. BL003A) was purchased from Biosharp. Tissue Mitochondria Isolation Kit (#C3606) and Cell Mitochondria Isolation Kit (#C3601) were purchased from Beyotime. The One‐Step Terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) Apoptosis Assay Kit (#C1089) was purchased from Beyotime.

Wang M., Su L., Sun J., Cai L., Li X., Zhu X., Song L., Li J., Tong S., He Q., Cai M., Yang L., Chen Y., Wang L, & Huang X. (2022). FGF21 attenuates pulmonary arterial hypertension via downregulation of miR‐130, which targets PPARγ. Journal of Cellular and Molecular Medicine, 26(4), 1034-1049.

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MTT Assay for Cell Viability

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3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was purchased from MP (Solon, OH, USA). Dulbecco’s modified Eagle’s medium (DMEM), penicillin–streptomycin, phosphate-buffered saline (PBS), fetal bovine serum (FBS), Pierce BCA protein assay kit, and Super Signal™ West Pico Chemiluminescent Substrate kit were obtained from Thermo Fisher Scientific (Waltham, MA, USA). Calcein/propidium iodide (PI) kit, RIPA buffer, crystal violet, 4% paraformaldehyde, and PI/RNase kit were from Beyotime Biotechnology (Jiangsu, China). Trizol buffer was purchased from Invitrogen (Carlsbad, CA, USA). The primary antibodies against cyclin D1, c-Myc, PI3K, P-PI3K, Akt, P-Akt, HIF-1A, VEGFA, MAPK, and P-MAPK were purchased from Affinity (Cincinnati, OH, USA), and β-actin and GAPDH were from Cell Signaling Technology (Beverly, MA, USA). The secondary antibodies were obtained from Millipore Corporation (Temecula, CA, USA).

Zhang W., Sang S., Peng C., Li G.Q., Ou L., Feng Z., Zou Y., Yuan Y, & Yao M. (2022). Network Pharmacology and Transcriptomic Sequencing Analyses Reveal the Molecular Mechanism of Sanguisorba officinalis Against Colorectal Cancer. Frontiers in Oncology, 12, 807718.

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