Surface expression of neutrophil granule proteins was measured as in (42 (link)), using phycoerythrin (PE)-CD63 for primary granules and allophycocyanin (APC)-CD66b for secondary granules (or isotype controls PE-IgG1 and APC-IgM) (BioLegend). Data were acquired using a Cytek Northern Lights spectral flow cytometer and analyzed using FCS Express (De Novo Software). The median fluorescence of each sample was normalized to unstimulated neutrophils as negative control.
Pe cd63
Manufactured by BioLegend
Sourced in United States
PE)-CD63 is a fluorescently-labeled antibody that binds to the CD63 cell surface protein. CD63 is a member of the tetraspanin family and is commonly used as a marker for lysosomal-associated membrane proteins and exosomes.
Automatically generated - may contain errors
Lab products found in correlation
Cd66b apc, by BioLegend (3 mentions)
Igg1 pe, by BioLegend (2 mentions)
Igm apc, by BioLegend (2 mentions)
Fcs express, by De Novo Software (2 mentions)
Cd45 fitc, by BioLegend (1 mentions)
Cd29 fitc, by Thermo Fisher Scientific (1 mentions)
Cd90 fitc, by BioLegend (1 mentions)
Cd34 fitc, by BioLegend (1 mentions)
Cd73 fitc, by BioLegend (1 mentions)
Bovine serum albumin (bsa), by Thermo Fisher Scientific (1 mentions)
Lsrfortessa, by BD (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Glutamine, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Gemini Bio (1 mentions)
Biotin anti human cd63 antibody, by BioLegend (1 mentions)
Anti human cd63 antibody, by BioLegend (1 mentions)
Hepes buffer, by R&D Systems (1 mentions)
6 protocols using pe cd63
1
Neutrophil Granule Protein Expression
2
Granule-specific marker expression in neutrophils
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Surface presentation of granule-specific markers was measured by flow cytometry essentially as described in Ragland et al. (2017) (link). Briefly, adherent neutrophils exposed to Gc were lifted with 5 mM EDTA, washed with DPBS containing 0.1% dextrose, and simultaneously stained with PE-CD63 (Biolegend) and APC-CD66b (Biolegend) as indicators of primary and secondary granule exocytosis, respectively, or respective isotype controls (Biolegend PE-IgG1, κ and Biolegend APC-IgM, κ). Data were acquired using a FACSCalibur Benchtop Analyzer and analyzed using FlowJo software. The geometric means of fluorescence intensity for PE and APC were calculated from a gate that includes all granulocytes by side scatter and forward scatter.
3
Neutrophil Granule Protein Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Surface expression of neutrophil granule proteins was measured as in reference (42 (link)), using phycoerythrin (PE)-CD63 for primary granules and allophycocyanin (APC)-CD66b for secondary granules (or isotype controls PE-IgG1 and APC-IgM) (BioLegend). Data were acquired using a Cytek Northern Lights spectral flow cytometer and analyzed using FCS Express (De Novo Software). The median fluorescence of each sample was normalized to unstimulated neutrophils as negative control.
4
Characterizing MenSCs and Their Mitochondria
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
MenSCs or MenSC-derived mitochondria were stained and labeled with anti-human fluorophore-conjugated antibodies for CD73-FITC (BioLegend, Inc.; Cat# 344015), CD90-FITC(BioLegend, Inc. Cat#328107), CD34-FITC (BioLegend, Inc. Cat# 343503), CD45-FITC (BioLegend, Inc. Cat# 304005), CD63-PE (BioLegend, Inc. Cat# 353003) and CD29-FITC (eBioscience; Thermo Fisher Scientific, Inc. Cat# 11029942) and detected by flow cytometric analysis. Briefly, trypsinized cells or mitochondria were washed then re-suspended in ice-cold PBS containing 1% BSA (Invitrogen; Thermo Fisher Scientific, Inc.). Fluorophore-conjugated antibodies were added at concentrations recommended by the manufacturer's protocols (95 µl staining buffer, 5 µl fluorophore-conjugated antibodies) and incubated in the dark for 30 min, 25˚C. The cells or mitochondria were washed twice in staining buffer (eBioscience, Thermo Fisher Scientific, Inc.) and analyzed under a flow cytometer (LSR Fortessa; BD Biosciences). Data analysis was performed using FlowJo (BD Biosciences. v.7.6.5).
5
Jurkat and JinB8 T Cell Culture
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
WT Jurkat and JinB8 T cells [14 (link)] were cultured using RPM1 1640 medium supplemented with 10% FBS (Gemini Bio, West Sacramento, CA, USA), glutamine and penicillin/streptomycin (Thermo Fischer Scientific, Waltham, MA, USA) as described previously [15 (link)]. Purified anti-human HLA-C Antibody, antibody, anti-human CD63 antibody, Biotin anti-human CD63 antibody, biotin anti-human HLA-A, B, C antibody, MHC-I-APC/640, CD63-PE and CD47-FITC antibodies were purchased from BioLegend, San Diego, CA, USA.
6
Basophil Activation Assay for Allergen Response
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
PBMCs isolated from non-allergic donors (5 10^5 cells) were stripped in 2 mL ice cold lactic acid buffer (0.13 M KCl, 0.05 M NaCl, 0.01 M lactic acid, pH = 3.9) for 30 s as described before [18 (link)]. After washing 3 times by PBS, cells were pre-incubated with sera from HDM-allergic individuals for 1 h at 37 °C. And then, cells were stimulated by different concentrations of synthetic mugwort tropomyosin or Derp 10 (50, 500 ng/mL) in hepes buffer containing IL-3 (R&D, Minneapolis, Minnesota, USA). In the meanwhile, cells exposed to FLMP (Sigma, St. Louis, USA)) were taken as a positive control. The reaction was stopped by EDTA buffer (20 mM). In the end, PBMCs were stained with basophil surface markers: CD123BV650, CCR3-APC-fire750 and CD63-PE (BioLegend, San Diego, CA, USA), and the percentages of CD63+CD123+CCR3+ cells were analysed by Flowjo software.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!