Histogel specimen processing gel

Manufactured by Thermo Fisher Scientific

Sourced in United States

HistoGel Specimen Processing Gel is a laboratory product designed to process and handle tissue specimens. It is a gelling agent used to encapsulate and support tissue samples during various laboratory procedures.

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Lab products found in correlation

4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions) Excelsior es tissue processor, by Thermo Fisher Scientific (1 mentions) Nunclon sphera, by Thermo Fisher Scientific (1 mentions) Neutral buffered formalin, by Avantor (1 mentions) Axioobserver z1 inverted microscope, by Hamamatsu Photonics (1 mentions) Normal donkey serum (nds), by Jackson ImmunoResearch (1 mentions) Citrate buffer ph 6, by Vector Laboratories (1 mentions) Prolong gold, by Thermo Fisher Scientific (1 mentions) Triton x 100, by Merck Group (1 mentions) Orca flash 4.0 camera, by Hamamatsu Photonics (1 mentions) Chromogranin a cga, by Abcam (1 mentions) Glycergel, by Agilent Technologies (1 mentions) Ab16667, by Abcam (1 mentions) Decloaking chamber, by Biocare Medical (1 mentions) Axio imager a2 microscope, by Zeiss (1 mentions) Deadend fluorometric tunel system, by Promega (1 mentions) Axiocam 503 mono, by Zeiss (1 mentions) Axio observer z1 microscope, by Zeiss (1 mentions) P4707, by Merck Group (1 mentions) Lsm 880 fluorescent microscope, by Zeiss (1 mentions)

8 protocols using histogel specimen processing gel

3D Culture Systems for Cancer Cell Spheroids

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Two 3D culture systems, AlgiMatrix (alginate scaffold six‐well plates; Cat. A10982‐02) and the Nunclon Sphera (scaffold‐free T75 flasks; Cat. 174952), were purchased from Thermo Fisher Scientific (Waltham, MA). HT‐29 cells were seeded at 2 × 10⁵ per well on Algimatrix plates and grown for 12 days for aggregate formation before harvesting. NCI‐H2228, NCI‐H1975, and KM12 cells were seeded at 5 to 6 × 10⁶ per Nunclon Sphera T75 flask and grown for 4 days for spheroid formation before harvesting. Spheroids and aggregates were processed in 10% neutral‐buffered formalin and then encapsulated in Richard‐Allen Scientific HistoGel Specimen Processing Gel (Thermo Fisher Scientific; Cat. HG‐4000‐012). HistoGel‐encapsulated samples were fixed overnight in Excelsior ES Tissue Processor (Thermo Fisher Scientific) followed by paraffin embedding on a Sakura Tissue‐Tek TEC Console (VWR Scientific, Radnor).

Bhaumik S., Boyer J., Banerjee C., Clark S., Sebastiao N., Vela E, & Towne P. (2020). Fluorescent multiplexing of 3D spheroids: Analysis of biomarkers using automated immunohistochemistry staining platform and multispectral imaging. Journal of Cellular Biochemistry, 121(12), 4974-4990.

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Immunohistochemical Analysis of Microtissues

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Microtissues were fixed in 10% Neutral Buffered Formalin (VWR, Radnor, PA, USA) for 1 hr at room temperature and embedded in HistoGel Specimen Processing Gel (ThermoFisher, Waltham, MA, USA) prior to paraffin processing. Five micron sections were cut and adhered to positively charged glass slides. Slides were deparaffinized with xylene and re-hydrated through a series of decreasing ethanol concentrations (100%, 100%, 95%, 80%, 70%). Epitope retrieval was performed by submersing slides in Citrate Buffer pH 6.0 (Vector Laboratories, Burlingame, CA, USA) in a 95°C water bath for 35 min. Slides were cooled at room temperature for 20 min and washed with PBS. Samples were permeabilized in 0.2% Triton X-100 (Sigma-Aldrich, St. Louis, MO, USA) for 5 min, blocked in 1.5% normal donkey serum (Jackson Immunoresearch, Westgrove, PA, USA) for 1 hr, and probed with primary and secondary antibodies (1:400) against cTnT and VIM and counterstained with Hoechst (1:10000) (antibody information in Data File S3). Coverslips were mounted with anti-fade mounting medium (ProlongGold, Life Technologies, Carlsbad, CA, USA) and samples were imaged on a Zeiss Axio Observer Z1 inverted microscope equipped with a Hamamatsu ORCA-Flash 4.0 camera.

Floy M.E., Givens S.E., Matthys O.B., Mateyka T.D., Kerr C.M., Steinberg A.B., Silva A.C., Zhang J., Mei Y., Ogle B.M., McDevitt T.C., Kamp T.J, & Palecek S.P. (2021). Developmental lineage of human pluripotent stem cell-derived cardiac fibroblasts affects their functional phenotype. FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 35(9), e21799.

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Immunostaining of Colonoid Cultures

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Colonoids were collected for immunostaining at the end of the experiments (Figure 1A). In brief, colonoids from each experimental condition were pooled (n=3-6 wells) and resuspended in 50 μL Richard-Allan Scientific™ HistoGel™ Specimen Processing Gel (#HG-4000-012, Thermo Fisher Scientific, Waltham, MA), then fixed in 10% buffered formalin for 24-48 hours before they were embedded in paraffin as described elsewhere (42 (link)). Formalin-fixed paraffin-embedded sections were deparaffinized, and antigen retrieval was accomplished by boiling the sections in citrate buffer (pH 6.0) or Tris-EDTA buffer (pH 9.0) for 15 minutes in a commercial microwave oven. The sections were blocked with tris-buffered saline (TBS) + 5% bovine serum albumin (BSA) for five minutes. Primary antibodies against KI67 (Dako Agilent, Santa Clara, CA), Cytokeratin 20 (CK20) (Dako Agilent), Mucin-2 (MUC2) (Abcam, Cambridge, Great Britain), and Chromogranin A (CGA) (Abcam) were diluted in TBS + Tween^® 20 + 1% BSA and incubated overnight at 4°C (SF 1, sheet 2). The secondary staining was performed with the MaxFluor immunofluorescence detection system (MaxVision Bioscience, Ontario, Canada). Lastly, the sections were counterstained with DAPI (Thermo Scientific) and mounted with Glycergel (Dako Agilent).

Walaas G.A., Gopalakrishnan S., Bakke I., Skovdahl H.K., Flatberg A., Østvik A.E., Sandvik A.K, & Bruland T. (2023). Physiological hypoxia improves growth and functional differentiation of human intestinal epithelial organoids. Frontiers in Immunology, 14, 1095812.

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Evaluating Histological Responses of D. medinensis to FBZ

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To evaluate histological changes in D. medinensis worms in response to FBZ treatment, 2.0- to 2.5-cm segments were selected randomly from five female D. medinensis worms isolated from FBZ-treated ferrets, the control ferret, and an additional five worms from untreated animals from previous work.¹⁹ (link) Sections were fixed and stored in 10% formol (3.7% formaldehyde), then embedded using HistoGel specimen processing gel (Thermo Scientific, Waltham, MA) allowing for the production of transverse sections through the worm segments.²⁰ Sections were cut from three levels (∼100 μm apart) of each worm segment and stained with hematoxylin and eosin. Each section was first assessed for histological integrity before scoring for any damage or degeneration in any of the major anatomic components of worms, including uterine contents.

Cleveland C.A., Garrett K.B., Box E.K., Thompson A.T., Haynes E.K., Elder D.L., Richards R.L., Majewska A.A., Guagliardo S.A., Wiegand R.E., Bryan II J.A., Torres-Velez F., Unterwegner K., Romero M., Zirimwabagabo H., Sidouin M., Oaukou P.T., Ada M.M., Ngandolo B.N., Mackenzie C.D., Geary T.G., Weiss A.J, & Yabsley M.J. (2022). Investigating Flubendazole as an Anthelmintic Treatment for Guinea Worm (Dracunculus medinensis): Clinical Trials in Laboratory-Reared Ferrets and Domestic Dogs in Chad. The American Journal of Tropical Medicine and Hygiene, 106(5), 1456-1465.

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Paraffin Sectioning and Immunohistochemistry of Organoids

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Tissue and organoids were processed for paraffin sectioning, and H&E staining was performed on 5-μm paraffin sections using standard protocols. Briefly, organoids were harvested with dispase II, washed in PBS, fixed in 4% paraformaldehyde at 4°C overnight, and centrifuged at 300 rcf for 5 min. The cell pellet was resuspended in 30 μl of HistoGel specimen processing gel (HG-4000-012, Thermo Fisher Scientific) before processing and embedding. For immunohistochemical staining, 5-μm paraffin sections were deparaffinized in xylene, rehydrated with a graded series of ethanol, and treated with a heat retrieval solution (RV1000M, Biocare Medical) in a digital electric pressure cooker (Decloaking Chamber, Biocare Medical). Slides were incubated with Ki-67 antibody (1:200 dilution; ab16667, Abcam) diluted in Da Vinci Green diluent (PD900M, Biocare Medical) overnight at 4°C. Slides were then incubated with a Polink-2 HRP Plus Rabbit DAB Detection System (D39-18, GBI Labs) according to the manufacturer’s instructions. Immunohistochemical staining of p53, PAX-8, and WT1 was performed by the Research Histology Core Laboratory at MD Anderson. Images were obtained with an Axio Imager A2 microscope (Carl Zeiss).

Zhang X., Yao J., Li X., Niu N., Liu Y., Hajek R.A., Peng G., Westin S., Sood A.K, & Liu J. (2023). Targeting polyploid giant cancer cells potentiates a therapeutic response and overcomes resistance to PARP inhibitors in ovarian cancer. Science Advances, 9(29), eadf7195.

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Apoptosis Detection in Multicellular Spheroids

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TUNEL stain was performed according to the manufacturer’s protocol (DeadEnd™ Fluorometric TUNEL System, Promega Corporation, Madison, WI)). Briefly, Spheres were collected and fixed in 4% paraformaldehyde, embedded in Histogel Specimen Processing Gel (Thermofisher Scientific, Waltham, MA) prior to embedding in paraffin. Spheres were sectioned at 5 μm each slice, deparaffinized using fresh xylene, rehydrated with 100, 95, 85, 70, 50% graded ethanol and washed with 0.85% NaCl at room temperature prior to fixing with 4% formaldehyde solution. Then the sphere sections were washed with PBS and pretreated with 20 µg/ml Proteinase K solution for 10 min, after which were washed with PBS prior to labeling with rTdT incubation better solution consisted of equilibrium buffer, nucleotide mix and rTdT enzyme as provided by the manufacturer. All sphere sections were counterstained with DAPI to visualize nuclei and apoptotic cells with fluorescein-12-dUTP fluorescein (green) were identified using Nikon Eclipse TE200 fluorescence microscope.

Tan J.J., Guyette J.P., Miki K., Xiao L., Kaur G., Wu T., Zhu L., Hansen K.J., Ling K.H., Milan D.J, & Ott H.C. (2021). Human iPS-derived pre-epicardial cells direct cardiomyocyte aggregation expansion and organization in vitro. Nature Communications, 12, 4997.

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Intestinal Organoid Histology and Staining

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Organoids were fixed in 5% neutral buffered formalin for 30 minutes, embedded in HistoGel specimen processing gel (HG-4000-012; ThermoFisher Scientific) before embedding in paraffin, and sectioned at 5 um. Intestinal alkaline phosphatase was detected using nitroblue tetrazolium chloride 5-bromo-4-chloro-3-indolyl phosphate (NBT/BCIP) according to published methods.⁵⁶ (link) For immunostaining on paraffin sections, slides were dewaxed in xylene, rehydrated, and heat-mediated antigen retrieval was conducted with citrate buffer (pH 6). For Goblet cells, sections were immunostained with mucin-2 primary antibody (sc-15334, 1:50 dilution; Santa Cruz Biotechnology, Dallas, TX). Secondary staining was conducted with Alexa 568 goat anti-rabbit IgG (A-11011, 1:1000 dilution; ThermoFisher Scientific). Samples were mounted in Vectashield with 4′,6-diamidino-2-phenylindole (H-1200; Vector Laboratories) to mark nuclei. Images were acquired using the Zeiss Axio Observer.Z1 microscope with Apotome, with an AxioCam 503 mono camera (Zeiss). Images were analyzed using ZEN software.

Lewis S.K., Nachun D., Martin M.G., Horvath S., Coppola G, & Jones D.L. (2019). DNA Methylation Analysis Validates Organoids as a Viable Model for Studying Human Intestinal Aging. Cellular and Molecular Gastroenterology and Hepatology, 9(3), 527-541.

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Immunofluorescence Staining of Organoids

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Organoids grown in Matrigel were initially fixed in 4% PFA (Millipore Sigma catalog no. 47608) for 1.5 h. After PBS washing, organoids were embedded in HistoGel Specimen Processing Gel (Thermo Fisher Scientific catalog no. HG-4000-012), processed with an automated tissue processor (Sakura Tissue-Tek VIP), and embedded into a paraffin block (Tissue-Tek TEC). Samples were sectioned at 4 μm onto poly-L-lysine coated slides (Sigma-Aldrich; catalog no. P4707) and air-dried at RT over-night for any subsequent immunofluorescence staining. All slides for fluorescence were deparaffinized and antigen retrieved in pH 6 citrate buffer for a total of 40 min. After protein blocking, nuclei were stained with DAPI (Abcam; catalog no. ab104139). Infiltrating PBMCs were pre-stained with Molecular Probes Vybrant CM-Dil as described above. Imaging was performed on a Zeiss LSM880 fluorescent microscope with Zen Black software.

Soldi R., Ghosh Halder T., Weston A., Thode T., Drenner K., Lewis R., Kaadige M.R., Srivastava S., Daniel Ampanattu S., Rodriguez del Villar R., Lang J., Vankayalapati H., Weissman B., Trent J.M., Hendricks W.P, & Sharma S. (2020). The novel reversible LSD1 inhibitor SP-2577 promotes anti-tumor immunity in SWItch/Sucrose-NonFermentable (SWI/SNF) complex mutated ovarian cancer. PLoS ONE, 15(7), e0235705.

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