Flow cytometry Anti-bodies Human - CD171/L1CAM

Flow cytometry is an immunophenotyping technique whereby sing cell suspensions are stained for either cell surface markers or intracellular proteins by fluorescently-labelled antibodies and analyzed with a flow cytometer, where fluorescently-labelled molecules are excited by the laser to emit light at varying wavelengths, which is then detected by the instrument. There are several key criteria which are required to be kept in mind while designing a flow experiment- 1. Antibody titration (optimal dilution of antibodies should be calculated in order to avoid over- or under- saturated signals for proper detection of surface and intracellular markers), 2. Precision (3 or more replicates of the sample should be used per experiment), 3. Specificity (proper isotype controls should be included in the experiment), 4. Day-to-day variability (experiments should be repeated 3 or more times to ensure consistency and avoid variability due to flow cytometer settings), 5. Antibody interaction (Fluorescence minus one or FMO should be used, which is the comparison of signals from panel minus one antibody vs. the full panel), and 6. Antibody stability (fluorescently-labelled antibodies should be stored at 4C).

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Found 2 matching solutions for this experiment

Anti-Human L1CAM Therapeutic Antibody Fab Fragment

Creative BioLabs

Upstream tips	Protocol tips	Downstream tips
	Cells were incubated with the human anti-L1CAM mAb, Ab417 (Cho et al., 2016), for 1 h at 4°C. After washing twice with phosphate buffered saline (PBS), cells were incubated with an anti-human IgG-FITC conjugate (Sigma) for 1 h at 4°C. After washing, propidium iodide (PI) was added to cells (Sigma), with PI-negative.	Cells were analyzed for bound antibody using a FACSCalibur flow cytometer (Becton-Dickinson, USA). Data acquisition and analyses were with the CellQuest software (Becton-Dickinson).

Protocol tips
Cells were incubated with the human anti-L1CAM mAb, Ab417 (Cho et al., 2016), for 1 h at 4°C. After washing twice with phosphate buffered saline (PBS), cells were incubated with an anti-human IgG-FITC conjugate (Sigma) for 1 h at 4°C. After washing, propidium iodide (PI) was added to cells (Sigma), with PI-negative.
Downstream tips
Cells were analyzed for bound antibody using a FACSCalibur flow cytometer (Becton-Dickinson, USA). Data acquisition and analyses were with the CellQuest software (Becton-Dickinson).

Manufacturer protocol Publication protocol 1 Relevant paper

CD171 (L1CAM) Antibody, anti-human, PE-Vio® 770, REAfinity™

Miltenyibiotec

Upstream tips	Protocol tips	Downstream tips
	>20 × 105 disaggregated sphere cells were stained with a stem cell‐specific antibody panel and analysed on a BDTM LSR II flow cytometer. To a volume of 100 μL cell suspension, 5 μL antibody mix was added, mixed and incubated for 20 minutes at 4°C in the dark and washed twice with 250 μL DPBS or FACS buffer.

Protocol tips
>20 × 105 disaggregated sphere cells were stained with a stem cell‐specific antibody panel and analysed on a BDTM LSR II flow cytometer. To a volume of 100 μL cell suspension, 5 μL antibody mix was added, mixed and incubated for 20 minutes at 4°C in the dark and washed twice with 250 μL DPBS or FACS buffer.

Manufacturer protocol Publication protocol 1 Relevant paper

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