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Alkaline phosphatase

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Alkaline Phosphatase is an enzymatic reagent used in various laboratory applications. It catalyzes the hydrolysis of phosphate esters in an alkaline environment. The core function of Alkaline Phosphatase is to facilitate biochemical reactions and analysis procedures that require the cleavage of phosphate groups from substrates.

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Lab products found in correlation

Alcian blue, by Thermo Fisher Scientific (1 mentions) Picrosirius red, by Thermo Fisher Scientific (1 mentions) Dpx solution, by Merck Group (1 mentions) Immpact dab, by Vector Laboratories (1 mentions) Axioplan 2 microscope, by Zeiss (1 mentions) Anti sox2, by Abcam (1 mentions) Anti oct4, by Abcam (1 mentions) Anti nanog, by Santa Cruz Biotechnology (1 mentions)

4 protocols using alkaline phosphatase

1

Tissue Histology and Staining Protocol

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Samples were fixed in 10% Neutralised Buffered Formalin (Sigma) at room temperature overnight, dehydrated in graded alcohols, paraffin embedded and sectioned at 4μm. Tissue slides were stained according to manufacturers’ instructions with Haematoxylin and Eosin (H&E) (Thermo Fisher), Alkaline Phosphatase (Vector Laboratories), Alcian-Blue Periodic Acid Schiff (Sigma). Picrosirius Red (PR), Elastic Van Gieson (EVG) and Alcian Blue (AB) (Thermo Fisher) staining was used to assess retention of collagen, elastin and glycosaminoglycans respectively.
Laween M., Isobel M., Sara C., Weston A.E., Riana G., Lucinda T., Peter F., Michael O., Anna K., Anna B., Laura N., Nikolaos A., Elizabeth H., Julia K., Tedeschi A.M., Pellegata A.F., Susanna E., Alessandro A.P., Lucy C., Nikhil T., Thomas G.M., Simon E., Paola B., De Coppi P, & Li V.S. (2020). Engineering transplantable jejunal mucosal grafts using patient-derived organoids from children with intestinal failure. Nature medicine, 26(10), 1593-1601.
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2

Immunohistochemical Staining Protocol

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Frozen sections were rinsed three times with 1× PBS for 10 min then fixed in 4% paraformaldehyde (PFA). The slides were washed three times with 1X PBS then incubated in 0.3% hydrogen peroxide for 10 min followed by 10 min incubation with 0.1% Triton × with three 1× PBS washes between the two incubations. The sections were blocked in 2% BSA for 45 min and then incubated with primary antibodies at 4 °C overnight. The following day, the slides were washed using 1× PBS and incubated with horseradish peroxidase or alkaline phosphatase enzyme tagged secondary antibody for 30 min. The corresponding substrate, ImmPACT DAB or alkaline phosphatase (Vector Laboratories) was then added, followed by counterstaining with methyl green or haematoxylin and mounting using a DPX solution (Sigma Aldrich).
Venugopal A., Michalczyk A., Khasraw M, & Ackland M.L. (2022). EMT Molecular Signatures of Pancreatic Neuroendocrine Neoplasms. International Journal of Molecular Sciences, 23(21), 13645.
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3

Histopathological Analysis of Mouse Knee Joints

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Formalin-fixed, EDTA-decalcified, paraffin-embedded mouse tissue specimens were sectioned (5 μm) and stained with either hematoxylin and eosin (H&E) or Safranin O. H&E-stained mouse knee joint sections were semiquantitatively evaluated for the following parameters by investigators who were blinded to animal genotype: inflammation (scale of 0–3), synovial hyperplasia (scale of 0–3), pannus (scale of 0–1), cartilage erosion (scale of 0–2), and bone loss (scale of 0–3). Scores were summed to determine the total histopathology index per knee per mouse (scale of 0–12). Representative fore paw and knee joint sections were also immunostained using rabbit anti-mouse fibrin(ogen) sera in conjunction with alkaline phosphatase (Vector Laboratories)/fast red visualization (Sigma) systems as previously described (6 (link)). All images were captured using a Zeiss Axioplan 2 microscope.
Raghu H., Jone A., Cruz C., Rewerts C.L., Frederick M.D., Thornton S., Degen J.L, & Flick M.J. (2014). Plasminogen Is a Joint-Specific Positive or Negative Determinant of Arthritis Pathogenesis in Mice. Arthritis & rheumatology (Hoboken, N.J.), 66(6), 1504-1516.
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4

Optimized iPS cell generation

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Llgl2 and Grb7 mRNA were cloned via Gateway Cloning into pMX retroviral plasmid backbone containing a T2A-mCherry in frame. Retroviral particles of the Llgl2 and Grb7 overexpression constructs along with pMX-human Oct4, Sox2, Klf4, and c-Myc (OSKM) were transfected into Plat-E retrovirus packaging cells. Viral supernatants were harvested 3 days post transfection and used freshly. 1 × 105 early passage MEFs were plated in 6-well plates 1 day before transducing twice by spinfection on subsequent days with viral supernatants and 8ug/ml of polybrene. MEFs transduced with OSKM vectors with or without Llgl2 or Grb7 vector were cultured onto gelatin-coated dish with ESC media for 7 or 14 days. iPS colonies emerged by day 14 and entire wells were stained with Alkaline Phosphatase (Vector labs) for quantification. Picked iPS colonies were expanded for 2 days before harvesting mRNA for qPCR quantification and western blot analysis using anti-OCT4 (1:1000, Abcam Ltd, Cambridge, MA), anti-SOX2 (1:1000, Abcam), and anti-NANOG (1:1000, Santa Cruz).
Moon B.S., Huang D., Gao F., Cai M., Lyu G., Zhang L., Chen J, & Lu W. (2023). Long range inter-chromosomal interaction of Oct4 distal enhancer loci regulates ESCs pluripotency. Cell Death Discovery, 9, 61.
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