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Ab68538

Manufactured by Abcam
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Ab68538 is a recombinant monoclonal antibody for the detection of Glutathione S-Transferase. The product is suitable for use in various immunoassay applications.

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Lab products found in correlation

Cm3050, by Leica (2 mentions) Alexa fluor 555 conjugated goat anti rabbit igg, by Thermo Fisher Scientific (2 mentions) Ab20920, by Abcam (2 mentions) Lsm 710, by Zeiss (1 mentions) Rat anti mouse cd11b, by BD (1 mentions) Rat anti mouse cd45, by BD (1 mentions) Rat anti mouse ly6g, by BD (1 mentions) Multiskan fc, by Thermo Fisher Scientific (1 mentions) A3687, by Merck Group (1 mentions) P nitrophenyl phosphate substrate solution, by Merck Group (1 mentions) Paraplast, by Leica (1 mentions) Aperio versa, by Leica (1 mentions) Ma5 14520, by Thermo Fisher Scientific (1 mentions) Pa5 16301, by Thermo Fisher Scientific (1 mentions) Ab5694, by Abcam (1 mentions)

5 protocols using ab68538

1

Immunohistochemistry of Lung Tissues for P. aeruginosa Detection

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Lungs were harvested at necropsy and fixed in 10% neutral buffered formalin for 16 h, washed with PBS and dehydrated in 70% ethanol and embedded in paraffin. Sections (5 µm) were cut using a Microm 355H microtome and stained with hematoxylin and eosin. For immunohistochemistry, sections were treated with 0.05% trypsin for 10 min at 37°C, blocked with goat serum and stained with rabbit anti-P. aeruginosa antibody (Abcam ab68538, Cambridge, MA, USA). Slides were washed, blocked with goat serum and then stained with AlexaFluor 555-conjugated goat anti-rabbit IgG (Thermo Scientific, A21428) and DAPI. For lung cryosection images, fluorescent microparticles were delivered and mice were immediately euthanized. Lungs were harvested and mounted in OCT embedding compound and flash frozen in liquid nitrogen. 10–20 µm thick sections were cut using a cryostat (Leica CM3050 S). The slides were dried at room temperature for 30 mins, washed with PBS and stained with DAPI.
Agarwal R., Johnson C.T., Imhoff B.R., Donlan R.M., McCarty N.A, & García A.J. (2018). Inhaled bacteriophage-loaded polymeric microparticles ameliorate acute lung infections. Nature biomedical engineering, 2(11), 841-849.
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2

Visualizing Bacterial Lung Infection and Immune Response

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Animals with pneumonia were intravenously injected with 100 nmol of FAM-labeled synthetic peptide, at 24–48 h
post-infection, and the peptide was allowed to circulate for 30–60 minutes. Following terminal cardiac perfusion with PBS,
lungs and other organs were isolated, fixed in 4% (w/v) paraformaldehyde (PFA) in PBS overnight, washed with PBS,
cryo-protected in sucrose solution [30% (w/v) in PBS] overnight, and processed through OCT embedding.
Ten-μm sections were cut and analyzed by fluorescence microscopy. The primary antibodies for immunofluorescence were
rabbit anti-Staphylococcus aureus, (ab20920, Abcam); rabbit anti-Fluorescein (A889, Invitrogen); rabbit
anti-Pseudomonas (ab68538, Abcam); rat anti-mouse CD11b (550282, BD Pharmingen); rat anti-mouse CD45 (550539,
BD Pharmingen); rat anti-mouse Ly-6G (551459, BD Pharmingen). These antibodies were incubated in diluted (1%) blocking
buffer overnight at dilutions 1:100 or 1:200 at 4°C, the sections were washed with PBS-T and incubated with secondary
antibodies diluted 1:500 or 1:1000, in 1% blocking buffer for one hour at room temperature. Subsequently sections were
washed with PBS-T, counterstained with DAPI (1ug/mL) in PBS for 10 minutes, washed with PBS, mounted using mounting media
(Molecular Probes, Life Technologies), and examined using a confocal microscope (Zeiss LSM-710).
Hussain S., Joo J., Kang J., Kim B., Braun G.B., She Z.G., Kim D., Mann A.P., Mölder T., Teesalu T., Carnazza S., Guglielmino S., Sailor M.J, & Ruoslahti E. (2018). Antibiotic-loaded nanoparticles targeted to the site of infection enhance antibacterial efficacy. Nature biomedical engineering, 2(2), 95-103.
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3

Quantifying Pseudomonas aeruginosa Antibodies

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Overnight P. aeruginosa cultures were centrifuged at 16,000 × g for 5 min at 4 °C, washed twice with PBS, and re-suspended in 4% paraformaldehyde (PFA, 15710-S, Electronic Microscopy Sciences, USA) in PBS for 30 min at 4 °C. After fixation, cultures were washed once with PBS, adjusted to 0.5 OD600nm in PBS, and pipetted onto Maxisorp plates (50 µl/well). After washing three times with PBS wells were blocked by adding 3% (w/v) bovine serum albumin (BSA) (80400-100, Alpha diagnostics, 50 µl/well) in PBS and then incubated with rabbit anti-P. aeruginosa polyclonal antibody (50 µl/well, ab68538, Abcam) diluted 1:1000 in PBS for 90 min at RT. After three washes in PBS, the plate was incubated with goat anti-rabbit IgG conjugated to alkaline phosphatase diluted 1:2000 (A3687, Sigma, 50 µl/well) in PBS for 1 h at RT. After three washes with AP buffer (100 mM Tris-HCl, 100 mM NaCl, 1 mM MgCl2, pH 9.5), 50 µl of p-nitrophenyl phosphate substrate solution (Sigma) were added to each well and incubated for 30-40 min at room temperature in the dark. Absorbance was measured at 405 nm using a Multiskan FC (Thermo Scientific).
Singh S., Almuhanna Y., Alshahrani M.Y., Lowman D.W., Rice P.J., Gell C., Ma Z., Graves B., Jackson D., Lee K., Juarez R., Koranteng J., Muntaka S., Daniel A. M.i.t.c.h.e.l.l., da Silva A.C., Hussain F., Yilmaz G., Mastrotto F., Irie Y., Williams P., Williams D.L., Cámara M, & Martinez-Pomares L. (2021). Carbohydrates from Pseudomonas aeruginosa biofilms interact with immune C-type lectins and interfere with their receptor function. NPJ Biofilms and Microbiomes, 7, 87.
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4

Immunohistochemistry of Lung Tissues for P. aeruginosa Detection

Check if the same lab product or an alternative is used in the 5 most similar protocols
Lungs were harvested at necropsy and fixed in 10% neutral buffered formalin for 16 h, washed with PBS and dehydrated in 70% ethanol and embedded in paraffin. Sections (5 µm) were cut using a Microm 355H microtome and stained with hematoxylin and eosin. For immunohistochemistry, sections were treated with 0.05% trypsin for 10 min at 37°C, blocked with goat serum and stained with rabbit anti-P. aeruginosa antibody (Abcam ab68538, Cambridge, MA, USA). Slides were washed, blocked with goat serum and then stained with AlexaFluor 555-conjugated goat anti-rabbit IgG (Thermo Scientific, A21428) and DAPI. For lung cryosection images, fluorescent microparticles were delivered and mice were immediately euthanized. Lungs were harvested and mounted in OCT embedding compound and flash frozen in liquid nitrogen. 10–20 µm thick sections were cut using a cryostat (Leica CM3050 S). The slides were dried at room temperature for 30 mins, washed with PBS and stained with DAPI.
Agarwal R., Johnson C.T., Imhoff B.R., Donlan R.M., McCarty N.A, & García A.J. (2018). Inhaled bacteriophage-loaded polymeric microparticles ameliorate acute lung infections. Nature biomedical engineering, 2(11), 841-849.
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5

Histological and Immunohistochemical Analysis of Tissue Scaffolds

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Samples of the scaffolds were fixed in 4% paraformaldehyde for 24 h and stored at 4 °C in 70% ethanol, dehydrated, and embedded in Paraplast (Leica Biosystems). For morphological analysis, 5 μm sections were stained with Hematoxylin–Eosin (H&E) and Masson’s trichrome. Stained tissue sections were scanned at × 40 equivalent resolution using a slide scanner Aperio Versa (Leica Biosystems) and images were captured with Aperio ImageScope 12.4.6 software. Immunohistochemistry was performed according to previously published protocols69 using rabbit polyclonal anti-CD31 (PA5-16301 Invitrogen), 1:50 dilution; rabbit polyclonal anti-α-SMA (ab5694 Abcam), 1:1000 dilution; rabbit monoclonal anti-ki67 (MA5-14520 Invitrogen), 1:50 dilution. Bacterial visualization was performed using rabbit polyclonal anti-Pseudomonas (Abcam ab68538), 1:1000 dilution; rabbit polyclonal anti-Staphylococcus (Abcam ab20920), 1:1000 dilution; and rabbit polyclonal anti-Enterococcus (Abcam ab19980), 1:1000 dilution.
Cárdenas-Calderón C., Veloso-Giménez V., González T., Wozniak A., García P., Martín S.S., Varas J.F., Carrasco-Wong I., Vera M, & Egaña J.T. (2022). Development of an implantable three-dimensional model of a functional pathogenic multispecies biofilm to study infected wounds. Scientific Reports, 12, 21846.
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