Annexin V assay was performed64 (link),65 (link) by first dissociating cortical organoids and monolayer neurons, which were subsequently resuspended in Annexin V binding buffer (Invitrogen) with Annexin V-CF488A conjugate (Biotium, Inc., Hayward, CA, USA) and Hoechst 33342 (Chemometec) and incubated for 15 min at 37°C. After a PBS wash, the cells were resuspended in Annexin V binding buffer (Invitrogen) supplemented by 10 μg/mL propidium iodide (Chemometec). Cells were loaded into NC-Slide A2 chambers and assessed with a Chemometec NucleoCounter NC-3000 cytometer using the preoptimized Annexin V assay.
Annexin 5 cf488a conjugate
Manufactured by Biotium
Sourced in United States
The Annexin V-CF488A conjugate is a fluorescent labeling reagent used for the detection and quantification of phosphatidylserine (PS) exposure on the cell surface. Annexin V is a calcium-dependent phospholipid-binding protein that has a high affinity for PS, and the CF488A fluorophore is attached to it, allowing for visualization and analysis of apoptotic cells.
Automatically generated - may contain errors
Lab products found in correlation
Hoechst 33342, by ChemoMetec (5 mentions)
Propidium iodide, by ChemoMetec (5 mentions)
Annexin 5 binding buffer, by Biotium (4 mentions)
Nc 3000 cell analyzer, by ChemoMetec (4 mentions)
Annexin 5 binding buffer, by Thermo Fisher Scientific (2 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
Cell proliferation reagent wst 1, by Roche (2 mentions)
Via1 cassette, by ChemoMetec (2 mentions)
4 6 diamidino 2 phenylindole (dapi), by ChemoMetec (2 mentions)
Nucleocounter nc 3000 cytometer, by ChemoMetec (1 mentions)
Trypsin edta solution, by Thermo Fisher Scientific (1 mentions)
Nc slides a8, by ChemoMetec (1 mentions)
Penicillin g, by Merck Group (1 mentions)
Nc slides a2 and a8, by ChemoMetec (1 mentions)
Eos rebel t3i eos 600d camera, by Canon (1 mentions)
Ecl western blotting substrate, by Thermo Fisher Scientific (1 mentions)
Polyvinylidene fluoride (pvdf), by Merck Group (1 mentions)
Tween 20, by Merck Group (1 mentions)
Amphotericin b, by Thermo Fisher Scientific (1 mentions)
Penicillin g, by Thermo Fisher Scientific (1 mentions)
11 protocols using annexin 5 cf488a conjugate
1
Annexin V Apoptosis Assay in Organoids
2
Neobavaisoflavone Sensitizes Anaplastic Astrocytoma Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human anaplastic astrocytoma cells (SW1783, HTB-13™) were obtained from ATCC (Manassas, VA, USA). Neobavaisoflavone (7-hydroxy-3-(4-hydroxy-3-(3-methyl-2-buten-1-yl)phenyl)-4H-1-benzopyran-4-one), penicillin G, and dimethyl sulfoxide (DMSO) were retrieved from Sigma-Aldrich Inc. (St. Louis, MO, USA). Dulbecco’s modified Eagle’s medium (DMEM) and fetal bovine serum (FBS) were acquired from Cytogen (Zgierz, Poland). Trypsin/EDTA solution was purchased from ThermoFisher Scientific (Waltham, MA, USA). Cell proliferation reagent WST-1 was obtained from Roche (Mannheim, Germany). NC-Slides™ A8 and Via1-Cassettes™, as well as Solution 3 (1 μg/mL DAPI, 0.1% Triton X-100 in PBS), Solution 7 (200 μg/mL JC-1 in DMSO), Solution 8 (1 μg/mL DAPI in PBS), Solution 15 (500 μg/mL Hoechst 33342, aqueous), Solution 16 (500 μg/mL propidium iodide, aqueous), were retrieved from ChemoMetec (Lillerød, Denmark). Neomycin sulfate was acquired from Amara (Kraków, Poland). Annexin V binding buffer and Annexin V-CF488A conjugate were obtained from Biotium (Fremont, CA, USA). In the study, the following drugs were used: doxorubicin (Doxorubicin Accord, Accord, Ahmedabad, India), etoposide (Etoposid-Ebewe, Ebewe Pharma, Ahmedabad, India), irinotecan (Irinotecan Accord, Accord, Ahmedabad, India). The rest of the chemicals were purchased from POCH S.A. (Gliwice, Poland).
3
Apoptosis Induction by Cell Cytometry
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Imaging cytometry was done on the NC-3000 cell analyzer (ChemoMetec, Allerod, Denmark) with reagents supplied by ChemoMetec. To determine induction of cell death, apoptotic cells were stained with AnnexinV-CF488A conjugate (Biotium, Fremont, CA, USA) in AnnexinV buffer and Hoechst 33342 (10 μg /mL) for 15 min at 37 °C, followed by several washes. Three independent assays with two technical replicates per dosage were performed. Data were analyzed on Graphpad Prism software using Mann–Whitney tests and are depicted as column graphs with average values and standard deviation.
4
Apoptosis and Cell Cycle Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Imaging cytometry was carried out on the NC-3000 cell analyzer (ChemoMetec, Allerod, Denmark) with reagents supplied by ChemoMetec. To determine the induction of cell death apoptotic cells were stained with AnnexinV-CF488A conjugate (Biotium, Fremont, CA, USA) in AnnexinV buffer and Hoechst 33,342 (10 μg/mL) for 15 min at 37 °C, followed by several washes. Propidium iodide was added shortly before imaging. For cell cycle analysis, cells were incubated in lysis buffer with DAPI (10 μg/mL) for 5 min at 37 °C before imaging on the NC-3000 cell analyzer.
5
Apoptosis Quantification in Cortical Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cortical organoids, primary fetal neurons, and astrocytes were manually dissociated and resuspended in Annexin V binding buffer (Invitrogen). Next, Annexin V-CF488A conjugate (Biotium, Inc., Hayward, CA, USA) was added, followed by Hoechst 33342 (Chemometec). After a PBS wash, the cells were resuspended in Annexin V binding buffer (Invitrogen) containing 10 μg/mL propidium iodide (Chemometec). Cells were loaded into NC‐Slide A2 chambers, and the “Annexin V Assay” was run with the NucleoCounter NC-3000.
6
Apoptosis and Cell Cycle Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Imaging cytometry was carried out on the NC-3000 cell analyzer (ChemoMetec, Allerod, Denmark) with reagents supplied by ChemoMetec. To determine the induction of apoptotis, cells were stained with AnnexinV-CF488A conjugate (Biotium, Fremont, CA, USA) in AnnexinV buffer and Hoechst 33,342 (10 μg/mL) for 15 min at 37 °C followed by several washes. Propidium iodide was added shortly before imaging. According to Annexin V and PI staining intensity, cells were classified as vital (Ann lo, PI lo), early apoptotic (Ann hi, PI lo), late apoptotic (Ann hi, PI hi), or necrotic (Ann lo, PI hi). For cell cycle analysis, cells were incubated in a lysis buffer with DAPI (10 μg/mL) for 5 min at 37 °C before imaging on the NC-3000 cell analyzer. According to DAPI staining, cell intensities were classified as subG1 (<2N), G0/G1 (2N), S phase (2–4N), or G2 phase (4N). Statistical analysis was performed using an unpaired t-test on GraphPad Prism 9 software. Data are depicted as column bar graphs with SD values.
7
Neobavaisoflavone Cytotoxicity Evaluation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Neobavaisoflavone (7-hydroxy-3-(4-hydroxy-3-(3-methyl-2-buten-1-yl)phenyl)-4H- -1-benzopyran-4-one), dimethyl sulfoxide (DMSO) and penicillin G were retrieved from Sigma-Aldrich Inc. (St. Louis, MO, USA). Gibco Astrocyte Medium (DMEM, N-2 Supplement, One Shot fetal bovine serum), Trypsin/EDTA solution were purchased from ThermoFisher Scientific (Waltham, MA, USA). Dulbecco’s modified Eagle’s medium (DMEM) and fetal bovine serum (FBS) were obtained from Cytogen (Zgierz, Poland). Via1-Cassettes™, NC-Slides™ A2 and A8, as well as Solution 3 (1 μg/mL DAPI, 0.1% Triton X-100 in PBS), Solution 7 (200 μg/mL JC-1), Solution 8 (1 μg/mL DAPI in PBS), Solution 15 (500 μg/mL Hoechst 33342), Solution 16 (500 μg/mL propidium iodide) were acquired from ChemoMetec (Lillerød, Denmark). Annexin V binding buffer and Annexin V-CF488A conjugate were purchased from Biotium (Fremont, CA, USA). Neomycin sulfate was retrieved from Amara (Kraków, Poland). In the study, the following drugs were used: etoposide (Etoposid-Ebewe, EbewePharma, Ahmedabad, India), and doxorubicin (Doxorubicin Accord, Accord, Ahmedabad, India). The rest of the chemicals were purchased from POCH S.A. (Gliwice, Poland).
8
Annexin V Assay for Cell Death
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were seeded on a 30mm glass botton dish (Cellvis, Mountain View, CA, USA) and grown for 2 days in DMEM+GFs. Cell death was assessed using Annexin V CF 488A conjugate (Biotium, Fremont, CA, USA) on unfixed cells according to the manufacturer's instructions. In brief, media was first removed and supernatant examined and substrate microscopically imaged. Cells on the substratum were then washed two times in 1X binding buffer (Biotium, Fremont, CA, USA). Cells were incubated in staining solution containing binding buffer and 1.25 μg/ml Annexin V conjugate for 30 min at room temperature. Stained cells were washed two times with binding buffer. Cells were imaged using a Nikon TE2000 inverted epifluorescent microsope (Nikon Instruments, Melville, NY, USA). Images were taken with an EOS Rebel T3i/EOS 600D camera (Canon, Lake Success, NY, USA) within 1 hour, as recommended by the manufacturer
9
Moxifloxacin Antibiotic Cytotoxicity Evaluation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Avelox solution for infusion (1 bottle of 250 ml containing 400 mg moxifloxacin as hydrochloride) was obtained from Bayer Healthcare Pharmaceuticals Inc. (Germany). Growth medium DMEM, penicillin G, amphotericin B, neomycin sulfate, fetal bovine serum (FBS), and trypsin/EDTA were purchased from Cytogen (Poland). Cell Proliferation Reagent WST-1 was produced by Roche GmbH (Germany). Staining reagents: DAPI (1 µg/ml), 0.1% Triton X-100 in PBS; VitaBright-48 (VB-48), propidium iodide (PI) and acridine orange (AO) solution; JC-1 (200 µg/ml in DMSO); Hoechst 33,342 (500 µg/ml), PI (500 µg/ml); NC-Slide A2 and A8; Via-1-Cassette were obtained from ChemoMetec (Denmark). Annexin V-CF488A conjugate and Annexin V binding buffer were obtained from Biotium Inc. (USA). Antibodies: anti-Mcl-1 (4572, polyclonal antibody that detects an endogenous level of human Mcl-1, source: rabbit), anti-MITF (D5G7V, monoclonal antibody that recognizes an endogenous level of total MITF protein, source: rabbit), and anti-GAPDH (14C10, rabbit monoclonal antibody) were obtained from Cell Signaling (USA). Anti-Rabbit IgG (A154), RIPA Buffer, Tween-20, and PVDF membranes were purchased from Sigma-Aldrich Inc. (USA). Pierce BCA Protein Assay Kit, penicillin G, amphotericin B, and ECL Western Blotting Substrate were purchased from Thermo Fisher Scientific (USA).
10
Apoptosis and Cell Cycle Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Imaging cytometry was carried out on an NC-3000 cell analyzer (ChemoMetec, Allerod, Denmark), with reagents supplied by ChemoMetec. To determine the induction of apoptosis, cells were stained with AnnexinV-CF488A conjugate (Biotium, Fremont, CA, USA) in AnnexinV buffer and Hoechst 33,342 (10 μg/mL) for 15 min at 37 °C, followed by several washes. Propidium iodide was added shortly before imaging. According to Annexin V and PI staining intensity, cells were classified as vital (Ann lo, PI lo), early apoptotic (Ann hi, PI lo), late apoptotic (Ann hi, PI hi) or necrotic (Ann lo, PI hi). For cell cycle analysis, cells were incubated in lysis buffer with DAPI (10 μg/mL) for 5 min at 37 C before imaging on the NC-3000 cell analyzer. According to DAPI staining intensity, cells were classified as subG1 (<2 N), G0/G1 (2 N), S phase (2–4 N), or G2 phase (4 N). Statistical analysis was performed using the unpaired t-test on GraphPad Prism version 10. Data are depicted as stacked column bar graphs.
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!