To mark PCs, brain sections were incubated with a rabbit anti-calbindin D28K antibody (1:1000, PA1-931, Invitrogen, Waltham, MA) at 4 °C for 24–48 h. After being washed in PBS, sections were incubated with a goat anti-rabbit Cy5 (1:500, A10523, Invitrogen) at room temperature for 3 h. After another rinse with PBS, sections were mounted on slides and cover-slipped with a mounting medium (H-1500, Vector Labs). To identify subtypes of c-Fos positive neurons following the social recognition test, sections were incubated with primary antibodies of guinea pig anti-c-Fos (1:1000, 226308, Synaptic Systems, Gottingen, Germany), rabbit anti-CaMKII (1:200, PA5-99558, Invitrogen) and mouse anti-GAD67 (1:500, MAB5406, Millipore, Burlington, MA) at 4 °C for 24–48 h. After being washed in PBS, sections were incubated with secondary antibodies of goat anti-guinea pig Alexa 488 (1:1000, A11073, Invitrogen), goat anti-rabbit Alexa 555 (1:1000, A21428, Invitrogen) and goat anti-mouse Alexa 647 (1:1000, A21236, Invitrogen) at room temperature for 3 h. After another rinse with PBS, sections were mounted on slides and cover-slipped with a mounting medium (H-1500, Vector Labs). Images were taken with a Zeiss LSM 710 confocal microscope and Zen 2.0 software (Oberkochen, Germany).
A11073
Manufactured by Thermo Fisher Scientific
Sourced in United States
The A11073 is a laboratory instrument designed for the analysis and detection of various analytes. It utilizes a specific analytical technique to provide accurate and reliable measurements. The core function of this product is to facilitate precise quantitative and qualitative analysis within a laboratory setting.
Automatically generated - may contain errors
Lab products found in correlation
A11001, by Thermo Fisher Scientific (4 mentions)
A21428, by Thermo Fisher Scientific (4 mentions)
A31573, by Thermo Fisher Scientific (4 mentions)
Ab7842, by Abcam (4 mentions)
Ab5905, by Merck Group (3 mentions)
A10523, by Thermo Fisher Scientific (2 mentions)
A32732, by Thermo Fisher Scientific (2 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (2 mentions)
Ab184955, by Abcam (2 mentions)
A31570, by Thermo Fisher Scientific (2 mentions)
Prolong gold, by Thermo Fisher Scientific (2 mentions)
A11012, by Thermo Fisher Scientific (2 mentions)
A 11076, by Thermo Fisher Scientific (2 mentions)
A11011, by Thermo Fisher Scientific (2 mentions)
A10042, by Thermo Fisher Scientific (2 mentions)
Hoechst 33342, by Thermo Fisher Scientific (2 mentions)
Lsm 710, by Zeiss (2 mentions)
Mab5406, by Merck Group (1 mentions)
A21236, by Thermo Fisher Scientific (1 mentions)
H 1500, by Vector Laboratories (1 mentions)
32 protocols using a11073
1
Immunohistochemical Analysis of Neuronal Subtypes
2
Immunofluorescent Staining of Kidney Tissue
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Double-immunofluorescent staining was performed on the sections of frozen mouse kidney tissue. The sections were incubated with rabbit anti-P2X7R antibody (1:200, Alomone, APR-004), rabbit anti-NLRP3 antibody (1:200, NOVUS, NBP-12448), and Guinea pig anti-nephrin antibody (1:200, PROPEN, GP-N2) for overnight at 4°C. After washing, the sections were incubated with Alexa 555 labeled goat anti-rabbit secondary antibody (1:50, Invitrogen, A-32732) and Alexa 488 labeled goat anti-Guinea pig secondary antibody (1:50, Invitrogen, A-11073) for 30 min at 37°C. After washing, the sections were mounted with a mounting solution containing DAPI and then observed with a confocal laser scanning microscope (Leica TSC SP5II, Germany).[21 (link)]
3
Immunostaining of Frozen Tissue Sections
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For immunofluorescence staining, frozen tissue sections were cut at 7-8 μm, air dried and permeabilized with ice-cold methanol for 15 min. Sections were briefly washed twice with PBS and placed in 0.3% Triton X-100 10 min before blocking with 10% KPL (5140-0011, SeraCare) in PBS for 1 h at room temperature. The sections were incubated with primary antibodies against phospho-Stat3 (1:200; 9145, Cell Signaling Technology), insulin (1:200; PA1-26938, ThermoFisher), proinsulin (1:200; MAB-13361, R&D), Reelin (1:1000; AF3820, R&D), and GFAP (1:200; LS-B4775, LifeSpan Bioscience) at 4 °C overnight. Sections were washed three times with PBS and incubated for 1 h in the dark with the corresponding fluorophore-conjugated secondary antibodies, Alexa Fluor 488 Chicken anti-rabbit IgG (1:500; A-21441, Invitrogen), Alexa Fluor 488 Goat anti-guinea pig IgG (1:1000; A-11073, Invitrogen), Alexa Fluor 555 Goat anti-rabbit IgG (1:1000; A-32732, Invitrogen), Alexa Fluor 555 Goat anti-chicken IgG (1:500; A-21437, Invitrogen), Alexa Fluor 647 Goat anti-mouse IgG (1:500; A-21235, Invitrogen). Sections were counterstained using DAPI (1:1000 from 1 mg/ml; D9542; Sigma) to visualize cell nuclei and cover slipped with Fluoroshield (Sigma, F6182). Images were captured using the Olympus confocal microscope FV3000. ImageJ software was used to quantify MFI per islet area in the Il22ra1 x Ins2-cre animals.
4
SARS-CoV-2 Immunohistochemistry in Lung Tissue
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5μm sections of formalin-fixed, paraffin-embedded lung were mounted on charged glass slides, baked for one hour at 60°C, and passed through Xylene, graded ethanol, and double distilled water to remove paraffin and rehydrate tissue sections. A microwave was used for heat induced epitope retrieval. Slides were heated in a high pH solution (Vector Labs H-3301), rinsed in hot water and transferred to a heated low pH solution (Vector Labs H-3300) where they were allowed to cool to room temperature. Sections were washed in a solution of phosphate-buffered saline and fish gelatin (PBS-FSG) and transferred to a humidified chamber, for staining at room temperature. Tissues were blocked with 10% normal goat serum (NGS) for 40 minutes, followed by a 60-minute incubation with a guinea pig anti-SARS antibody (BEI NR-10361) diluted 1:1000 in NGS. Slides were washed and transferred to the humidified chamber for a 40-minute incubation with a goat anti-guinea pig secondary antibody (Invitrogen A11073) tagged with Alexa Fluor 488 and diluted 1:1000 in NGS. Following washes, DAPI (4’,6-diamidino-2-phenylindole) was used to label the nuclei of each section. Slides were mounted using a homemade anti-quenching mounting media containing Mowiol (Calbiochem#475904) and DABCO (Sigma #D2522) and imaged at 20X with a Zeiss Axio Slide Scanner.
5
Microglial and Astrocyte Labeling
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The same protocol previously described was used, applying the specific antibodies. Primary antibodies used were rabbit anti-Iba-1 (1:1000, 019-19741, Wako) and guinea pig anti-GFAP (1:1000, 173 004, Synaptic system) to label microglial cells and astrocytes, respectively. The secondary antibodies employed were AlexaFluor-594 (1:500, A-11012, Invitrogen) for Iba-1 and AlexaFluor-488 (1:500, A-11073, Invitrogen) for GFAP labeling.
6
Immunostaining of Skin Whole Mounts
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Wound samples were isolated, and subcutaneous fat was removed by scraping. Tissues were fixed in periodate-lysine-4% paraformaldehyde (PLP) buffer overnight at 4°C. Whole skin samples were permeabilized and blocked for 3 h in 5% BSA/5% NGS/1% Triton X in PBS and subsequently stained with primary antibodies (anti-K14 [PRB155P; Covance] and anti-K10 [GP-K10; Progen]) in blocking solution overnight at RT. After three washes with 0.2% Tween-20 in PBS, secondary antibodies (goat anti-guinea pig AF488 [A11073; Invitrogen] and chicken anti-rabbit AF647 [A21443; Invitrogen]) were incubated for 6 h at RT. Skin samples were washed three times with 0.2% Tween-20 in PBS and mounted in VectaShield Hard Set. All whole mounts were imaged with an inverted Leica SP8 Dive system with an Insight X3 laser (Spectra-Physics). Different fluorophores were excited as follows: CFP with MP 840 nm, AF 488 at 488 nm, and Alexa 647 at 633 nm. CFP was collected at 430–470 nm, AF 488 at 492–530 nm, and AF 647 at 640–700 nm. All images were collected in 12 bit with 25× water immersion objective (HC FLUOTAR L N.A. 0.95 W VISIR 0.17 FWD 2.4 mm).
7
Immunostaining of Retinal Cell Markers
8
Immunofluorescence and Western Blot Antibody Protocols
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The following antibodies were used for immunofluorescence: guinea pig anti-insulin (A056401, Dako, 1:1000), rabbit anti-phospho-CREB (Ser133) (9198, Cell Signaling, 1:250), rabbit anti-Oxytocin (T-4084, Peninsula Laboratories, 1:1000), mouse anti-V5 tag (R960-25, Invitrogen, 1:250), goat anti-guinea pig conjugated to Alexa Fluor 488 (A11073, Invitrogen, 1:500), goat anti-rabbit conjugated to Cy5 (A10523, Invitrogen, 1:500), and goat anti-mouse conjugated to Alexa647 (115-605-003, Jackson ImmunoResearch, 1:500). The following antibodies were used for Western blot: mouse anti-V5 tag (R960-25, Invitrogen, 1:5000), goat anti-IgSF8 (AF3117, R&D Systems, 1:500), mouse anti-Alpha-synuclein (610786, BD Biosciences, 1:500), rabbit anti-Beta-arrestin (ab32099, Abcam, 1:1000), rabbit anti-c-Raf (53745, Cell Signalling, 1:500), rabbit anti-GAPDH (2118, Cell Signalling, 1:1000), goat anti-rabbit conjugated to HRP (401393, Sigma, 1:10,000), rabbit anti-goat conjugated to HRP (401515, Sigma, 1:10,000), and goat anti-mouse conjugated to HRP (401253, Sigma, 1:10,000).
9
Immunofluorescence Labeling of Brain Sections
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For immunofluorescence, brain sections were blocked for 1 h with QuickBlockTM blocking buffer for Immunol labeling (P0260, Beyotime, CHN) and then incubated overnight at 4°C with primary antibody diluted in the blocking solution (rabbit anti‐IBA1, 1:500, PTR2404; mouse anti‐CD68, 1:100, ab955, Abcam49 ; guinea pig anti‐vGluT1, 1:1000, ab5905, Millipore, mouse anti‐GAD65, 1:50, ab_528264, Developmental Studies Hybridoma Bank). After washing, the sections were incubated with corresponding secondary antibodies: Alexa 488‐conjugated goat anti‐mouse IgG (1:1000, A21121, Invitrogen), Alexa 488‐conjugated goat anti‐guinea pig IgG (1:1000, A11073, Invitrogen), Alexa 568‐conjugated goat anti‐mouse IgG (1:1000, A21134, Invitrogen) or Alexa 647‐conjugated goat anti‐rabbit IgG (1:1000, A32733, Invitrogen) for 2 h at 37°C temperature; then slices were mounted on glass coverslips with DAPI Fluoromount‐G (0100–20, Southern Biotech). The sections were imaged by a laser scanning spectral confocal microscope (TCS SP8, Leica Microsystems).
10
Multiprotein Imaging in Brain Tissue
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After the selection of DG- and BLA-containing tissue sections, washings with PBS and antigen retrieval technique proceedings were carried out as described previously. Unspecific bindings were blocked with BSA solution (3% in 0.3% of Triton-X-100 in PBS for 90 min, RT). Sections were incubated for 48 h at 4 °C with gentle agitation with guinea pig polyclonal anti Arc (156 004; 1:1500; Synaptic Systems, Goettingen, Germany), mouse monoclonal anti GluN1 antibody (32-0500; 1:100; Thermo Fischer Scientific) and rabbit monoclonal anti Homer1 (ab184955; 1:150; Abcam). Later, brain tissue samples were incubated with the following secondary antibodies: anti-mouse conjugated with Alexa Fluor 555 (A-31570, 1:1000, Invitrogen), anti-guinea pig conjugated with Alexa 488 (A-11073, 1:1000, Invitrogen), and anti-rabbit conjugated with Alexa 647 (A-31573; 1:1000, Invitrogen) for 4 h, as described in the previous section. Sections were also incubated with DAPI (1:25,000) for 1 min and mounted with ProLong Gold (Invitrogen) in gelatinized slides.
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