Histoclear hs200

Manufactured by National Diagnostics

Sourced in United States

Histoclear (HS200) is a laboratory reagent used in the processing and preparation of tissue samples for microscopic examination. It serves as a clearing agent, facilitating the removal of paraffin or other embedding media from tissue sections prior to staining and analysis. The core function of Histoclear is to provide a clear, transparent medium for the examination of tissue samples.

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Lab products found in correlation

Clarion mounting medium, by Merck Group (2 mentions) S0809, by Agilent Technologies (1 mentions) Prolong gold antifade, by Thermo Fisher Scientific (1 mentions) Mkb 2213, by Vector Laboratories (1 mentions) X0909, by Agilent Technologies (1 mentions) Paraformaldehyde (pfa), by Euromedex (1 mentions) Hm340e, by Thermo Fisher Scientific (1 mentions) Cytoseal, by Thermo Fisher Scientific (1 mentions) Eukitt mounting medium, by Merck Group (1 mentions) Dab substrate kit, by Abcam (1 mentions) Celltracker green cmfda, by Thermo Fisher Scientific (1 mentions) Axiovision 4, by Zeiss (1 mentions) Ethidium homodimer 1, by Thermo Fisher Scientific (1 mentions) Axiovert 200 microscope, by Zeiss (1 mentions) Dab kit, by Thermo Fisher Scientific (1 mentions)

6 protocols using histoclear hs200

Immunofluorescence Staining of Paraffin Sections

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Paraffin sections were warmed on a heating block set at 65°C for 10 minutes to facilitate bonding of the tissue with the glass slide. Tissue then was deparaffinized in Histo-Clear (HS-200; National Diagnostics, Atlanta, GA) and a series of ethanol, followed by rehydration and antigen retrieval in citrate buffer (S1699; Dako) in a pressure cooker set for high pressure for 15 minutes. After antigen retrieval, slides were cooled on ice and blocked at room temperature for 1.5 hours with serum-free protein block (X0909; Dako). Mouse on mouse block (MKB-2213; Vector Laboratories, Burlingame, CA) was used for mouse antibodies following the manufacturer’s instructions. Primary antibodies were diluted in antibody diluent with background reducing components (S3022; National Diagnostics) and were incubated overnight at 4°C in a humidified chamber. After 3 washes in PBS for 5 minutes, sections were incubated with secondary antibodies diluted 1:200 in antibody diluent (S0809; Dako, Santa Clara, CA) for 1 hour at room temperature. Hoechst (1:1000 in PBS, 62249, 12.3 mg/mL; Thermo Fisher Scientific) was added to stain the nuclei for 5 minutes. Sections then were washed 3 times in PBS for 5 minutes each. Coverslips were added to sections with ProLong Gold Antifade (P36934; Thermo Fisher Scientific) and allowed to dry before imaging.

Engevik A.C., Krystofiak E.S., Kaji I., Meyer A.R., Weis V.G., Goldstein A., Coutts A.W., Melkamu T., Saqui-Salces M, & Goldenring J.R. (2021). Recruitment of Polarity Complexes and Tight Junction Proteins to the Site of Apical Bulk Endocytosis. Cellular and Molecular Gastroenterology and Hepatology, 12(1), 59-80.

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Histological Analysis of Mouse Epididymis

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One caput epididymis and one testis per animal were fixed in 4% paraformaldehyde (Euromedex, Souffelweyersheim, France) for 24 h and 48 h, respectively. Tissues were then dehydrated in an increasing ethanol gradient and placed in Histoclear (HS200; National Diagnostics, Atlanta, GA, USA) for 2 h before paraffin embedding. Five µm-thick sections were made using a microtome (HM340E; Thermo Fisher Scientific, Illkirch, France). Tissue sections were then deparaffinized by passage through Histoclear and progressively rehydrated in a decreasing ethanol gradient. Masson’s Trichrome staining was performed using an automated deparaffinization and staining machine (HMS 70, Microm) and the slides were finally mounted with Cytoseal (Thermo Fisher Scientific, Illkirch, France). The photomicrographs presented for epididymal histology are focused on the caput segment 1/segment 2 boundary. This is because this region of the mouse epididymis caput has notable characteristics (in terms of, among others, tubule size/shape, presence of stereocilia and spermatozoa in the lumen of the tubules, cell distribution, etc.). Moreover, segment 1 of the mouse caput is highly irrigated by blood vessels, which makes it the most exposed segment to any circulating molecule.

Hug E., Villeneuve P., Bravard S., Chorfa A., Damon-Soubeyrand C., Somkuti S.G., Moazamian A., Aitken R.J., Gharagozloo P., Drevet J.R, & Saez F. (2023). Loss of Nuclear/DNA Integrity in Mouse Epididymal Spermatozoa after Short-Term Exposure to Low Doses of Dibutyl Phthalate or Bisphenol AF and Its Mitigation by Oral Antioxidant Supplementation. Antioxidants, 12(5), 1046.

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Immunohistochemical Analysis of Bladder Tissue

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Human bladder TMAs (T121a, T122, T123, T124, T125, HBlaU066Su01) were purchased from US Biomax, Rockville, MD 20849, US. Normal urinary bladder tissue sections (BLA01) were obtained from Pantomics, Inc 2600 Hilltop Drive, Suite C130, BLDG B, Richmond, CA 94806. Rabbit monoclonal anti-TM [EPR4051] antibody (AB108189), HistoReveal (AB103720) and DAB substrate kit (AB64238) were purchased from Abcam, Cambridge Science Park Milton Rd, Milton, Cambridge CB4 0FL, UK. Clearing reagent HistoClear (HS-200) was obtained from National Diagnostics, Atlanta, Georgia, US. Bovine serum albumin (BSA) (A4503), goat anti-rabbit HRP (A0545) conjugate, absolute ethanol (24,103), Triton X-100 (T9284–500mL), 30% hydrogen peroxide (95321), Eukitt® mounting medium (03989–100mL), and counterstains Harris hematoxylin (HHS32) and eosin (HT110116) were obtained from Sigma, Poole, UK. Phosphate buffered saline (10x) (AM9624) was obtained from Invitrogen by Thermo Fisher Scientific Baltics UAB, V. A. Graiciuno 8, Vilnius, LT-02241. All other reagents, unless otherwise indicated, were obtained from Sigma, Poole, UK.

Watt J., Maguire D.G., Reid C.N., Lamont J.V., Fitzgerald S.P, & Ruddock M.W. (2020). Thrombomodulin Expression in Bladder Cancer Tissue and Its Association with Prognosis and Patient Survival. Research and Reports in Urology, 12, 157-165.

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Cell Viability Assay with Fluorescent Dyes

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Metabolically active and necrotic cells were labelled with 10 µg/ml Cell Tracker Green CMFDA (5-chloromethylfluorescein diacetate, C7025; Life Technologies) and 5 µg/ml Ethidium Homodimer-1 (E1169; Life Technologies), respectively, for an hour. The constructs were incubated in tissue culture medium for 45 minutes and then fixed with 4% paraformaldehyde in PBS. The specimens were embedded in paraffin wax and sequential sections were cut in sections of 7 μm. After de-waxing with Histoclear (HS200; National Diagnostics, Leicestershire, UK) and re-hydration through graded ethanol, cell nuclei were stained with DAPI (4’,6-diamidino-2-phenylindole, dilactate, D3571; Life Technologies). Fluorescent images were acquired with Axiovert 200 microscope (Carl Zeiss Microscopy, Jena, Germany) equipped with Zeiss AxioCam HR colour camera, AxioCam MR3 monochrome camera and the Zeiss AxioVision 4.6 software.

Ching K.Y., Andriotis O., Sengers B, & Stolz M. (2021). Genipin crosslinked chitosan/PEO nanofibrous scaffolds exhibiting an improved microenvironment for the regeneration of articular cartilage. Journal of Biomaterials Applications, 36(3), 503-516.

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Hematoxylin and Eosin (H&E) Staining Protocol

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H&E staining was performed on cell aggregate frozen sections as indicated by the manufacturer (Richard-Allen Scientific, MI, USA), using hematoxylin (7211, Richard-Allen Scientific, MI, USA) and eosin staining (7111, Richard-Allen Scientific, MI, USA). The slides were then cleared 3 times in 100% ethanol. Slides were then washed 3 times using Histo-Clear (HS-200, National Diagnostics, NC, USA) for 5 min, after which the slides were mounted using Clarion mounting medium (CO487, Sigma-Aldrich, MO, USA). Slides were then cured overnight at 37 °C before imaging.

Valdoz J.C., Franks N.A., Cribbs C.G., Jacobs D.J., Dodson E.L., Knight C.J., Poulson P.D., Garfield S.R., Johnson B.C., Hemeyer B.M., Sudo M.T., Saunooke J.A., Kartchner B.C., Saxton A., Vallecillo-Zuniga M.L., Santos M., Chamberlain B., Christensen K.A., Nordin G.P., Narayanan A.S., Raghu G, & Van Ry P.M. (2022). Soluble ECM promotes organotypic formation in lung alveolar model. Biomaterials, 283, 121464.

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Immunohistochemical Staining Protocol

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Cell aggregate sections were permeabilized using 0.1% Triton X-100 for 10 min and then blocked using Seablock buffer for 1hr at room temperature. Primary antibodies (Table S4) were diluted in 10% Seablock in PBST and incubated overnight at 4 °C. After intensive washing, HRP-conjugated secondary antibodies were then diluted to 1:500 in 10% Seablock PBST and incubated for an hour at RT. The chromogen was developed using a DAB kit (34002, Thermo Scientific, MO, USA) for 15 min at RT. Counterstaining for the nucleus was done using hematoxylin staining and bluing reagent. The slides were then cleared 3 times in 100% ethanol. Slides were then washed 3 times using Histo-Clear (HS-200, National Diagnostics, NC, USA) for 5 min, after which the slides were mounted using Clarion mounting medium (CO487, Sigma Aldrich, MO, USA). Slides were then cured overnight at 37 °C before imaging.

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