Journal: Oncology Research

Article Title: RNA Interference of Biot2 Induces G 1 Phase Arrest and Apoptosis in Mouse Colorectal Cancer Cell Line

doi: 10.3727/096504014X14146137738583

Figure Lengend Snippet: Biot2 genes downregulated arrest of CT26 cell cycle. The effect of Biot2-shRNA on inducing G 1 cell cycle arrest in CT26 cells. After treatment with Biot2-shRNA for 48 h, (A) the cell cycle distributions were determined by flow cytometry. (B) The protein expressions of cycle regulatory proteins and cyclin-dependent kinase cyclin D1, CDK2 and (C) the expression of several CKIs, p16, p21, and p27, were measured by Western blotting.

Article Snippet: Rabbit anti-CDK2 (1:1,000), rabbit anti-p16 (1:1,000), rabbit anti-p27 (1:1,000), and mouse anti-PCNA (1:1,000) were purchased from BOSTER (China).

Techniques: shRNA, Flow Cytometry, Expressing, Western Blot

Journal:

Article Title: Loss of function of the tumor suppressor DKC1 perturbs p27 translation control and contributes to pituitary tumorigenesis

doi: 10.1158/0008-5472.CAN-09-4730

Figure Lengend Snippet: A) Chromatographs showing a novel mutation in the DKC1 locus identified in a human pituitary adenoma (left). Scheme of the mutation, an A1493G variation located on one DKC1 allele leads to the amino acid change S485G in the C-terminal portion of the protein (right). B) Analysis of DKC1S485G protein stability. Densitometric analysis of the DKC1 levels was done at each time point after blockage of protein synthesis as indicated. GFP was co-transfected for transfection efficiency and β-actin was used as loading control. C) DKC1 protein (left) and mRNA (right) levels in a normal pituitary and in the tumor bearing the mutation DKC1S485G. Densitometric analysis of DKC1 over β-actin protein levels in each sample is shown (bottom left).

Article Snippet: Supernatants were used for Western blot analysis by standard protocols using anti-p27 and anti-DKC1 antibodies (clone DCS-72; MBL international, Woburn, MA, USA, Atlas antibodies Stockholm, Sweden).

Techniques: Mutagenesis, Transfection, Control

Journal: Developmental biology

Article Title: Temporal and Spatial Effects of Sonic Hedgehog Signaling in Chick Eye Morphogenesis

doi: 10.1006/dbio.2000.0195

Figure Lengend Snippet: Viral-mediated Shh expression and anti-Shh antibody production by 5E1 hybridoma cells. (A–F′) Transverse sections of stage 24 eyes infected by viruses at stage 10 are shown. (A) and (B) show sections derived from a Shh virus-infected eye hybridized for Shh and Pct1 antisense probes, respectively. Note that the uneven ectopic expression of Shh mRNA (A) leads to a more evenly distributed Ptc1 mRNA in the adjacent section (B). (C) and (D) show the same cross section derived from a Shh virus-infected eye stained for the anti-viral antibody P27 (C, red) and the anti-Shh antibody 5E1 (D, green), respectively. (E) and (F) show the same section from a control virus RCAS-infected eye stained with P27 (E, red) and 5E1 antibodies (F, green), respectively. (F9) shows an enlarged view of the central retina in (F). Both Shh (C) and control RCAS (E) virus-infected eyes show positive staining for the viral GAG protein; however, Shh virus-infected eyes show Shh protein signals (D) elevated above the endogenous Shh protein staining signals detected in the ganglion cell layer (arrow, F9). (G–L) Sections of stage 27 embryos injected at stage 17 with anti-Shh 5E1 hybridoma cells (H, J, L) or control 3C2 hybridoma cells (G, I, K). All sections were incubated with anti-mouse secondary antibodies (green). 5E1 hybridoma cell-injected embryos show secondary antibody labeling of Shh expressing cells in the zona limitans (H, arrows) and in the ventral midline (J, arrow) of the diencephalon, as well as of 5E1 hybridoma cells in the vitreal space of the eye (L, arrowhead). Production of the control 3C2 antibody is also detected by the secondary antibody (K, arrowhead); however, no labeling of Shh producing cells is detected in the diencephalon (G, I). These data indicate that anti-Shh antibodies are produced by 5E1 cells in vivo and they diffuse and bind to endogenous Shh producing cells. Scale bars in (A–L) represent 100 μm; the scale bar in (F9) represents 25 μm. di, diencephalon; le, lens; pe, pigmented epithelium; os, optic stalk; zl, zona limitans.

Article Snippet: The anti-viral protein polyclonal antibody P27 was purchased from SPAFAS.

Techniques: Expressing, Infection, Derivative Assay, Virus, Staining, Control, Injection, Incubation, Antibody Labeling, Labeling, Produced, In Vivo

Journal: Frontiers in Microbiology

Article Title: Improved Detection of HIV Gag p24 Protein Using a Combined Immunoprecipitation and Digital ELISA Method

doi: 10.3389/fmicb.2021.636703

Figure Lengend Snippet: IP-Simoa assay for SIV p27 protein measurement. (A) SIV p27 Simoa assay standard curve. Linear signal response for recombinant p27 from 0 to 28 pg/mL. (B) Recombinant p27 spiked into uninfected rhesus monkey rectal biopsy (prepared by soaking in lysis buffer) and run with IP Simoa showed enriched p27 signal proportional to the sample volume reduction (20-fold) and expected reduction of signal upon dilution indicating assay specificity. (C) p27 was enriched significantly from the lysate of viremic, SIV + monkey rectal biopsy ( p < 0.05, n = 4). No detectable p27 was observed in the flow through after IP demonstrating high efficiency of p27 IP from biopsy lysate matrix. See Materials and Methods for statistical analysis.

Article Snippet: For p27 Simoa after IP and elution, the same kit for p24 was used except that the detection antibody was replaced with biotin-labeled anti-p27 antibody from ABL Inc. (Rockville, MD, Cat# ABL-4324) instead of biotin anti-p24 antibody in the p24 kit.

Techniques: Recombinant, Lysis