Immpact vip substrate

Manufactured by Vector Laboratories

Sourced in United States

ImmPACT® VIP Substrate is a chromogenic substrate for use in immunohistochemistry and immunocytochemistry applications. It produces a purple-colored reaction product when catalyzed by horseradish peroxidase (HRP) or alkaline phosphatase (AP) enzyme labels.

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Lab products found in correlation

Ab272420, by Abcam (1 mentions) Ab150686, by Abcam (1 mentions) Decloaking chamber dc2012, by Biocare Medical (1 mentions) Methyl green, by Vector Laboratories (1 mentions) Horse serum, by Vector Laboratories (1 mentions) Citrisolv, by Thermo Fisher Scientific (1 mentions) Anti vwf antibody, by Merck Group (1 mentions)

3 protocols using immpact vip substrate

SARS-CoV-2 Spike Glycoprotein IHC in Lung Autopsy

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We selected autopsy lung samples from a cohort of 3 patient groups, which were diagnosed as ARDS, COVID-19, and influenza a virus subtype H1N1 infection (Table 2). Briefly, 4 μm thin FFPE sections were prepared and deparaffinized in xylene, followed by rehydration with a concentration gradient of ethanol. Tissue sections were then subjected to heat-induced epitope retrieval with Tris-EDTA buffer (pH 9) and quenched with 3% H₂O₂. The slides were then incubated with primary antibody against SARS spike glycoprotein (1:1000; Abcam, Cambridge, UK; Ab272420) in 1% BSA/PBS solution. The remaining IHC steps were performed as previously described [23 (link)], and the staining was developed with ImmPACT^® VIP Substrate (Vector Labs, Burlingame, CA, USA).

Wong D.W., Klinkhammer B.M., Djudjaj S., Villwock S., Timm M.C., Buhl E.M., Wucherpfennig S., Cacchi C., Braunschweig T., Knüchel-Clarke R., Jonigk D., Werlein C., Bülow R.D., Dahl E., von Stillfried S, & Boor P. (2021). Multisystemic Cellular Tropism of SARS-CoV-2 in Autopsies of COVID-19 Patients. Cells, 10(8), 1900.

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Tissue Preparation and Immunohistochemistry Protocol

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All tissue samples were FFPE, sectioned (3 to 4 μm thick), and dried for 1 hour at 65°C. Next, tissue samples were subjected to deparaffinization, rehydration, and heat-induced epitope retrieval using a Biocare Decloaking Chamber (DC2012) at 110°C for 7 min in the corresponding unmasking solution. Sections were then incubated with primary antibody overnight at 4°C, washed, and HRP-coupled secondary antibody for 30 min. Reagents were incubated at room temperature in a humidified slide chamber. Revelation was performed using ImmPACT VIP Substrate (SK-4605, Vectorlabs). All the stainings were counterstained with hematoxylin. For Masson’s trichrome, staining was performed according to manufacturer’s recommendations (ab150686, Abcam).

Samain R., Maiques O., Monger J., Lam H., Candido J., George S., Ferrari N., KohIhammer L., Lunetto S., Varela A., Orgaz J.L., Vilardell F., Olsina J.J., Matias-Guiu X., Sarker D., Biddle A., Balkwill F.R., Eyles J., Wilkinson R.W., Kocher H.M., Calvo F., Wells C.M, & Sanz-Moreno V. (2023). CD73 controls Myosin II–driven invasion, metastasis, and immunosuppression in amoeboid pancreatic cancer cells. Science Advances, 9(42), eadi0244.

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Quantifying lung vasculature and muscularization

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Using a Leica Microtome RM2255 (Leica Biosystems, Buffalo Grove, IL), we made formalin-fixed and paraffin-embedded 5 μm slices of left lungs, deparaffinized with CitriSolv (Fisher Scientific, Pittsburgh, PA), and then rehydrated with a series of gradient ethanol. Sections were boiled in antigen unmasking solution; blocked using 10% horse serum (Vector Laboratories, Burlingame, CA); incubated overnight first with a mouse monoclonal α-smooth muscle actin (α-SMA) antibody, clone 1A4 (Sigma-Aldrich, St. Louis, MO), and then with peroxidase-conjugated anti-mouse/anti-rabbit secondary antibody (RTU Vectastatin Universal, Vector Laboratories, Burlingame, CA); and finally stained with peroxidase sensitive ImmPACT diaaminobenzidine substrate (Vector Laboratories, Burlingame, CA). Similarly, the von-Willebrand factor (vWF) was stained with rabbit polyclonal anti-vWF antibody (Sigma-Aldrich, St. Louis, MO) and peroxidase sensitive ImmPACT VIP substrate (Vector Laboratories, Burlingame, CA).^{32 (link)} The nuclei were counterstained with methyl green (Vector Laboratories, Burlingame, CA), and slides were viewed at 200× using the Olympus IX81 microscope (Olympus Scientific Solutions Americas Corp., Waltham, MA). ImageJ software analyzed the images, determined the degree of muscularization, and measured the medial wall thickness (MWT).

Rashid J., Nahar K., Raut S., Keshavarz A, & Ahsan F. (2018). Fasudil and DETA NONOate, Loaded in a Peptide-Modified Liposomal Carrier, Slow PAH Progression upon Pulmonary Delivery. Molecular pharmaceutics, 15(5), 1755-1765.

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