Catalog #: NLLC1435R Datasheet / COA / SDS
50X Concentration, Azide Free
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All SSEA-4 Antibodies
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GloLIVE Human/Mouse SSEA-4 NL557 Live Cell Antibody Summary
Kit Summary
To detect SSEA-4 expression in live cells by immunocytochemical staining.
Why confirm marker expression in live cells before colony selection?
Maintaining pluripotent human embryonic stem cells and deriving induced pluripotent stem cells require colony selection and cell expansion. Picking colonies that contain pluripotent stem cells is typically achieved by manually analyzing colony morphology. Unfortunately, this critical process is time-consuming and labor-intensive.
In addition, morphology-based colony selection does not consistently guarantee pluripotent starting populations, since cells can fail to be fully reprogrammed or can naturally differentiate within a colony.
To address this need, R&D Systems offers GloLIVE™ antibodies that allow researchers to confirm stem cell marker expression in live cells before colony selection.
Live pluripotent stem cell imaging:
- Promotes the selection of high quality, undifferentiated stem cell colonies to reduce experimental variation.
- Verifies stem cell pluripotency in 30 minutes, ensuring efficient use of time and reagents.
- Allows for the continued culture of pluripotent cells with no adverse effects on cell proliferation or stemness following staining.
Antibody Components
- GloLIVE NL557-conjugated Mouse Anti-Human/Mouse SSEA-4 Monoclonal Antibody.
- Supplied as 50X concentration of antibody in 0.5 mL PBS
Azide free and endotoxin tested (= 5 EU/mL), this antibody can be used for 12 tests, if a staining volume of 2 mL is used.
Specifications
Shipping Conditions
The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Storage
Store the unopened product at 2 - 8° C. Do not use past expiration date. Protect from light.
Applications
Immunocytochemistry
Species
Human Mouse
Limitations
This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.
Product Datasheets
Scientific Data
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Detection of SSEA-4 in Live BG01V Human Embryonic Stem Cells. The stem cell marker SSEA-4 was visualized in live BG01V human embryonic stem cells using a GloLIVE NL557-conjugated Mouse Anti-Human/Mouse SSEA-4 Monoclonal Antibody (Catalog # NLLC1435R; red). The cells were counterstained with Hoechst 33342 (blue). Positive staining for SSEA-4 in human stem cells is an indicator of pluripotency.
Background: SSEA-4
SSEA-4 is expressed on the surface of human embryonal carcinoma (EC) cells (the pluripotent stem cells of teratocarcinomas), human embryonic germ cells (EG), and human embryonic stem cells (ES). Expression of SSEA-4 is down-regulated following differentiation of human EC cells. In contrast, the differentiation of murine EC and EScells may be accompanied by an increase in SSEA-4 expression (1-4).
- Shevinsky, L.H. et al. (1982) Cell 30:697.
- Kannagi, R. et al. (1983) EMBO J. 2:2355.
- Thomson, J.A. and J.S. Odorico (2000) Trends Biotechnol. 18:53.
- Draper, J.S. et al. (2002) J. Anat. 200:249.
Long Name
Stage-specific Embryonic Antigen-4
Alternate Names
SSEA4; SSEA-4; SSEA-4;stage-specific embryonic antigen 4
Related Research Areas
- Embryonic and Induced Pluripotent Stem Cell Markers
- Mesenchymal Stem Cell Markers
- Neural Stem Cell Markers
Assay Procedure
Refer to the product datasheet for complete product details.
Briefly, SSEA-4 expression can be verified in live cells prior to colony selection or experimentation using this simple procedure:
- Add the SSEA-4 antibody directly to the media of cultured cells
- Wash the cells after a 30 minute incubation
- Select for and expand SSEA-4-positive colonies
Precautions
Use aseptic technique and sterile culturing conditions to prevent contamination.
Reagents Provided
GloLIVE NL557-conjugated Mouse Anti-Human/Mouse SSEA-4 Monoclonal Antibody. (Catalog#NLLC1435R):
Other Supplies Required
Reagents
- Stem cell culture media
Materials
- Human or mouse pluripotent stem cells
- Pipettes and pipette tips
- Serological pipette
Equipment
- 37 °C and 5% CO2 incubator
- Fluorescence microscope
Live Cell Immunocytochemistry Procedure Overview
To ensure sterility of cultures, all steps should be performed under sterile conditions.
- Add fluorochrome-conjugated primary antibody in appropriate culture media to cells.
- Incubate for 30 minutes.
- Replace with fresh culture media.
- Visualize using a fluorescence microscope.
Note: Culture with GloLIVE antibodies does not appear to affect cell proliferation or stemness as assayed by proliferation curves for 3 days post-antibody incubation and expression levels of SSEA-4 and Oct-3/4.
FAQs
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What is the recommended starting confluence to observe staining?
The optimal confluence would depend on the purpose of the staining. If the plan is to use all the cells in the flask after they have proliferated, then one can wait for the cells to become confluent or almost confluent (80-90% confluent) and then check the staining. If the goal is to to check on the cells mid-way through their growth and there are sufficient cells in the flask, one could check the staining then as well. The staining is not dependent on how many cells are in the flask, as long as there are sufficient cells.
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