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Cytoseal

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Cytoseal is a mounting medium used to secure coverslips in place on microscope slides. It is a solvent-based, synthetic formulation that dries to form a clear, non-fluorescent, and permanent seal. Cytoseal is designed to provide a secure, long-lasting attachment for the coverslip while maintaining optical clarity.

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Lab products found in correlation

Dab solution, by Agilent Technologies (3 mentions) Dab sparkle enhancer, by Biocare Medical (3 mentions) Hrp anti mouse, by Agilent Technologies (2 mentions) Er α clone 6f11, by Leica (1 mentions) P her2, by Cell Signaling Technology (1 mentions) Aluminum potassium sulfate, by Merck Group (1 mentions) Carmine, by Merck Group (1 mentions) Carmine alum, by Merck Group (1 mentions) Citrisolv, by Avantor (1 mentions) Citrisolv, by Thermo Fisher Scientific (1 mentions) Hematoxylin, by Vector Laboratories (1 mentions) Aperio slide scanner, by Leica (1 mentions) Vina green, by Biocare Medical (1 mentions) Mach 2 rabbit hrp polymer, by Biocare Medical (1 mentions) Warp red, by Biocare Medical (1 mentions) 2 methylbutane, by Thermo Fisher Scientific (1 mentions) Triton x 100, by Thermo Fisher Scientific (1 mentions)

5 protocols using cytoseal

1

Immunohistochemical Analysis of Tissue Samples

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Tissue specimens were fixed in neutral formalin buffered saline (10%)
and embedded in paraffin. Hematoxylin and eosin staining was performed using
standard methods and tissue specimens from experimental animals were reviewed in
a blinded fashion by a clinical pathologist (M. M. I.) For Ki67 staining,
3–4 micron tissue sections were cut from paraffin blocks and baked
overnight in a dry slide incubator, then deparaffinized on a Shandon-Lipshaw
Varistain using a series of incubations in xylene, ethanol, then water. Antigen
retrieval was achieved by incubating slides in Tris-HCL 9.0 AR buffer in a T-FAL
OPTIMA pressure cooker. Slides were rinsed, then endogenous peroxidase was
blocked by immersing slides in 3% hydrogen peroxide for 5 minutes. Following
washes in nanopure water and TBS-20, primary antibody was applied at a dilution
of 1:200 (Ki67, MIB-1 Clone, Dako). Following primary antibody incubation,
slides were washed and then incubated with envision-labelled polymer-HRP
Anti-Mouse (Dako). Slides were washed, then DAB+ solution (DakoCytomation) was
added for a 15-minute incubation, after which slides were rinsed with nanopure
water. Chromogen signal was enhanced using DAB Sparkle Enhancer (Biocare).
Slides were washed, then counterstained with Harris Hematoxylin, dehydrated,
cleared, then mounted using Cytoseal (VWR).
Bader D.A., Hartig S.M., Putluri V., Foley C., Hamilton M.P., Smith E.A., Saha P.K., Panigrahi A., Walker C., Zong L., Martini-Stoica H., Chen R., Rajapakshe K., Coarfa C., Sreekumar A., Mitsiades N., Bankson J.A., Ittmann M.M., O’Malley B.W., Putluri N, & McGuire S.E. (2018). Mitochondrial pyruvate import is a metabolic vulnerability in androgen receptor-driven prostate cancer. Nature metabolism, 1(1), 70-85.
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2

Immunohistochemical Analysis of Tissue Samples

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Tissue specimens were fixed in neutral formalin buffered saline (10%)
and embedded in paraffin. Hematoxylin and eosin staining was performed using
standard methods and tissue specimens from experimental animals were reviewed in
a blinded fashion by a clinical pathologist (M. M. I.) For Ki67 staining,
3–4 micron tissue sections were cut from paraffin blocks and baked
overnight in a dry slide incubator, then deparaffinized on a Shandon-Lipshaw
Varistain using a series of incubations in xylene, ethanol, then water. Antigen
retrieval was achieved by incubating slides in Tris-HCL 9.0 AR buffer in a T-FAL
OPTIMA pressure cooker. Slides were rinsed, then endogenous peroxidase was
blocked by immersing slides in 3% hydrogen peroxide for 5 minutes. Following
washes in nanopure water and TBS-20, primary antibody was applied at a dilution
of 1:200 (Ki67, MIB-1 Clone, Dako). Following primary antibody incubation,
slides were washed and then incubated with envision-labelled polymer-HRP
Anti-Mouse (Dako). Slides were washed, then DAB+ solution (DakoCytomation) was
added for a 15-minute incubation, after which slides were rinsed with nanopure
water. Chromogen signal was enhanced using DAB Sparkle Enhancer (Biocare).
Slides were washed, then counterstained with Harris Hematoxylin, dehydrated,
cleared, then mounted using Cytoseal (VWR).
Bader D.A., Hartig S.M., Putluri V., Foley C., Hamilton M.P., Smith E.A., Saha P.K., Panigrahi A., Walker C., Zong L., Martini-Stoica H., Chen R., Rajapakshe K., Coarfa C., Sreekumar A., Mitsiades N., Bankson J.A., Ittmann M.M., O’Malley B.W., Putluri N, & McGuire S.E. (2018). Mitochondrial pyruvate import is a metabolic vulnerability in androgen receptor-driven prostate cancer. Nature metabolism, 1(1), 70-85.
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3

Quantifying p-HER2 and ER+ in Xenograft Tissues

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IHC staining was executed with the help of the Pathology Core at the Lester and Sue Smith Breast Center, Baylor College of Medicine. Tissue sections were incubated at 58°C overnight in a dry slide incubator and deparaffinized in xylene and graded alcohol washes. Antigen retrieval was performed in 0.1 M Tris-HCl pH 9.0 (p-HER2 and ERα), followed by quenching in 3% H2O2. The following antibodies were used to stain for 1 hour at room temperature: ERα (clone 6F11, Novocastra, 1:200) and p-HER2 (Cell Signaling, 2243L, 1:50). After washing in TBS, EnVision labeled polymer-HRP anti-mouse or anti-rabbit antibodies (Dako) were added for 30 minutes at room temperature. Slides were washed with TBS and then developed with DAB+ solution (Dako) and DAB sparkle enhancer (Biocare). After washing in TBS, slides were counterstained with hematoxylin, dehydrated, and cleared before coverslipping with Cytoseal (VWR). p-HER2 and ER+ stained cells were quantified in lung/ovary sections of L755S fat pad (FP) xenografts and ovary sections from HER2 WT and HER2 S310F or L755S MIND xenograft mice.
Kalra R., Chen C.H., Wang J., Salam A.B., Dobrolecki L.E., Lewis A., Sallas C., Yates C.C., Gutierrez C., Karanam B., Anurag M., Lim B., Ellis M.J, & Kavuri S.M. (2022). Poziotinib Inhibits HER2-Mutant–Driven Therapeutic Resistance and Multiorgan Metastasis in Breast Cancer. Cancer Research, 82(16), 2928-2939.
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4

Whole Mount Tissue Preparation

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X-gal stained whole mounts were post-fixed in 4% PFA, washed twice with 1X PBS, dehydrated through an increasing ethanol gradient, cleared of lipids in Carnoy’s Fixative (60% Ethanol, 30% Chloroform, 10% Glacial Acetic Acid) for 2 h, and further cleared in Citrisolv (Fisher Scientific, Suwanne, GA) for 2 h. Glands were pressed flat between the slide and coverslip under a heavy weight for 30 min, and imaged on a Leica dissecting microscope Model WILD M3Z (Leica Microsystems, Bannockburn, IL) with an Optronics digital camera Model 60800 (Goleta, CA). The glands were then re-hydrated through a decreasing ethanol gradient and counterstained with Carmine alum (500 mL distilled water containing 1 g Carmine and 2.5 g aluminum potassium sulfate; Sigma Aldrich, St Louis, MO) diluted 1:4 in distilled water. Glands were once again dehydrated in ethanol, cleared in Carnoy’s Fixative and Citrisolv, and pressed flat before mounting under a coverslip with Cytoseal (VWR, West Chester PA) then re-photographed.
Spina E., Simundza J., Incassati A., Chandramouli A., Kugler M.C., Lin Z., Khodadadi-Jamayran A., Watson C.J, & Cowin P. (2022). Gpr125 is a unifying hallmark of multiple mammary progenitors coupled to tumor latency. Nature Communications, 13, 1421.
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5

Immunohistochemical Analysis of Placental Tissues

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Human placental explants were fixed in 4% PFA and incubated overnight in 30% sucrose solution. Tissues were then flash frozen in OCT media (Sakura) using 2-methylbutane (ACROS Organics) cooled on dry ice. 20 µm slices were made from tissue blocks and affixed to slides in chilled acetone. Tissues were washed once in 1x TBS Buffer (Abcam) for 5min and permeabilized with 0.3% Triton-X 100 (Fisher Scientific) in TBS for 20min at RT. After washing 3 times in 1x TBS, antigen retrieval was performed by submerging tissue slides in 1x DIVA Decloaker solution (Biocare Medical) and boiling in vegetable steamer for 30min. Tissues were washed 3 times in 1x TBS and incubated with primary antibodies (made up in TBS with 1% BSA and 0.05% Tween-20) overnight at 4°C – Ms an ti-4G2 (3 ug/ml), Rb anti-CD163 (1:300; Abcam). Next, tissues were washed and incubated with either Mach 2 mouse AP polymer (Biocare Medical) or Mach 2 rabbit HRP polymer (Biocare Medical) for 30min at RT. Slides were washed and stained with Warp Red (for AP polymer; Biocare Medical) or Vina Green (for HRP polymer; Biocare Medical) for approximately 5min. Counterstain was performed with Hematoxylin (Vector Laboratories) for 30sec and washing in ddH2O. Slides were dried for 3min at 60°C and mounted w ith Cytoseal (VWR). Slides were imaged using an Aperio Slide Scanner (Leica Biosystems) at 40x magnification.
Zimmerman M.G., Quicke K.M., O’Neal J.T., Arora N., Machiah D., Priyamvada L., Kauffman R.C., Register E., Adekunle O., Swieboda D., Johnson E.L., Cordes S., Haddad L., Chakraborty R., Coyne C.B., Wrammert J, & Suthar M.S. (2018). Cross-reactive dengue virus antibodies augment Zika virus infection of human placental macrophages. Cell host & microbe, 24(5), 731-742.e6.
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