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17-10228 Sigma-AldrichLentiBrite™ Histone H2B-RFP Lentiviral Biosensor

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Key Applications: Transfection, Immunofluorescence, Immunocytochemistry

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17-10228

1 vial (minimum of 3 x 10E8 IFU/mL)

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Key Spec Table

Key Applications	Detection Methods
TFX, IF, ICC	Fluorescent

Description
Catalogue Number	17-10228
Trade Name	LentiBrite Chemicon
Description	LentiBrite™ Histone H2B-RFP Lentiviral Biosensor
Overview	*Read our application note in Nature Methods!* http://www.nature.com/app_notes/nmeth/2012/121007/pdf/an8620.pdf (Click Here!) Learn more about the advantages of our LentiBrite Lentiviral Biosensors! Click Here Biosensors can be used to detect the presence/absence of a particular protein as well as the subcellular location of that protein within the live state of a cell. Fluorescent tags are often desired as a means to visualize the protein of interest within a cell by either fluorescent microscopy or time-lapse video capture. Visualizing live cells without disruption allows researchers to observe cellular conditions in real time. Lentiviral vector systems are a popular research tool used to introduce gene products into cells. Lentiviral transfection has advantages over non-viral methods such as chemical-based transfection including higher-efficiency transfection of dividing and non-dividing cells, long-term stable expression of the transgene, and low immunogenicity. EMD Millipore is introducing LentiBrite™ Lentiviral Biosensors, a new suite of pre-packaged lentiviral particles encoding important and foundational proteins of autophagy, apoptosis, and cell structure for visualization under different cell/disease states in live cell and in vitro analysis. Pre-packaged, fluorescently-tagged with GFP & RFP Higher efficiency transfection as compared to traditional chemical-based and other non-viral-based transfection methods Ability to transfect dividing, non-dividing, and difficult-to-transfect cell types, such as primary cells or stem cells Non-disruptive towards cellular function EMD Millipore’s LentiBrite™ Histone H2B-RFP lentiviral particles provide bright fluorescence and precise localization to enable live cell analysis of chromosomal dynamics in difficult-to-transfect cell types.
Alternate Names	Histone H2B
Background Information	Chromatin, the higher order structure of DNA and nucleosomes, constitutes the majority of the nucleus of the eukaryotic cell. Changes in chromatin structure are the essence of many essential nuclear processes, including transcription, mitosis, meiosis, and apoptosis. The nucleosome comprises an octomer of four core histone proteins (H2A, H2B, H3 and H4). Genetic fusions between histone H2B and fluorescent proteins have been widely used in live cells to visualize the dynamics of chromosomal architecture during various processes. In addition to its utility for studying normal mitosis, histone H2B-GFP has been utilized to define mechanisms for asymmetric inheritance of oncogenic double minute chromosomes. Monitoring histone H2B-GFP also permits continuous analysis of chromosomal degradation during apoptosis. Use of histone H2B-GFP has recently been extended to live animals and high-throughput siRNA screens. EMD Millipore’s LentiBrite™ Histone H2B-RFP lentiviral particles provide bright fluorescence and precise localization to enable live cell analysis of chromosomal dynamics in difficult-to-transfect cell types.

References

Product Information
Components	Histone H2B-TagRFP Lentivirus: One vial containing 25 µL of lentiviral particles at a minimum of 3 x 10E8 infectious units (IFU) per mL. For lot-specific titer information, please see lot specific “Viral Titer” in the product specifications of the datasheet. Promoter EF-1 (Elongation Factor-1) Multiplicty of Infection (MOI) MOI = Ratio of # of infectious lentiviral particles (IFU) to # of cells being infected. Typical MOI values for high transduction efficiency and signal intensity are in the range of 20-40. For this target, some cell types may require lower MOIs (e.g., HT-1080, HeLa, human mesenchymal stem cells (HuMSC)), while others may require higher MOIs (e.g., human umbilical vein endothelial cells (HUVEC), U2OS). NOTE: MOI should be titrated and optimized by the end user for each cell type and lentiviral target to achieve desired transduction efficiency and signal intensity.
Detection method	Fluorescent
Quality Level	MQ100

Applications
Key Applications	Transfection Immunofluorescence Immunocytochemistry
Application Notes	Fluorescence microscopy imaging: (See Figure 1 in datasheet) HeLa cells were plated in a chamber slide and transduced with lentiviral particles at an MOI of 20 for 24 hours. After media replacement and 48 hours further incubation, cells were fixed with formaldehyde and mounted. Images were obtained by oil immersion wide-field fluorescence microscopy. The histone H2B-RFP signal shows interphase nuclei, with the lower nuclei connected by a strand representing a segregation defect. Additional cell types and Hard-to-transfect cell types: (See Figure 2 in datasheet) Primary HUVECs were transduced at an MOI of 20. The image depicts an irregular interphase nucleus. HT1080 were transduced at an MOI of 40 for 24 hours. The image depicts an interphase nucleus. Human mesenchymal stem cells (HuMSC) transduced with lentiviral particles at an MOI of 40. The cell is in anaphase and displays lagging chromosomes. Mitosis in Various Stages: (See Figure 3 in datasheet) U2OS cells were plated in chamber slides and transduced with lentiviral particles at an MOI of 20 for 24 hours. The images show cells in various stages of mitosis. For optimal fluorescent visualization, it is recommended to analyze the target expression level within 24-48 hrs after transfection/infection for optimal live cell analysis, as fluorescent intensity may dim over time, especially in difficult-to-transfect cell lines. Infected cells may be frozen down after successful transfection/infection and thawed in culture to retain positive fluorescent expression beyond 24-48 hrs. Length and intensity of fluorescent expression varies between cell lines. Higher MOIs may be required for difficult-to-transfect cell lines.

Applications

Key Applications

Transfection
Immunofluorescence
Immunocytochemistry

Application Notes

Fluorescence microscopy imaging:
(See Figure 1 in datasheet)
HeLa cells were plated in a chamber slide and transduced with lentiviral particles at an MOI of 20 for 24 hours. After media replacement and 48 hours further incubation, cells were fixed with formaldehyde and mounted. Images were obtained by oil immersion wide-field fluorescence microscopy. The histone H2B-RFP signal shows interphase nuclei, with the lower nuclei connected by a strand representing a segregation defect.

Additional cell types and Hard-to-transfect cell types:
(See Figure 2 in datasheet)
Primary HUVECs were transduced at an MOI of 20. The image depicts an irregular interphase nucleus. HT1080 were transduced at an MOI of 40 for 24 hours. The image depicts an interphase nucleus. Human mesenchymal stem cells (HuMSC) transduced with lentiviral particles at an MOI of 40. The cell is in anaphase and displays lagging chromosomes.

Mitosis in Various Stages:
(See Figure 3 in datasheet)
U2OS cells were plated in chamber slides and transduced with lentiviral particles at an MOI of 20 for 24 hours. The images show cells in various stages of mitosis.

For optimal fluorescent visualization, it is recommended to analyze the target expression level within 24-48 hrs after transfection/infection for optimal live cell analysis, as fluorescent intensity may dim over time, especially in difficult-to-transfect cell lines. Infected cells may be frozen down after successful transfection/infection and thawed in culture to retain positive fluorescent expression beyond 24-48 hrs. Length and intensity of fluorescent expression varies between cell lines. Higher MOIs may be required for difficult-to-transfect cell lines.

Biological Information
Gene Symbol	H2B
Purification Method	PEG precipitation
UniProt Number	Q8N257

Physicochemical Information

Dimensions

Materials Information

Toxicological Information

Safety Information according to GHS

Safety Information

Product Usage Statements
Quality Assurance	Evaluated by transduction of HT-1080 cells and fluorescent imaging performed for assessment of target localization and transduction efficiency.
Usage Statement	This product contains genetically modified organisms (GMO). Within the EU GMOs are regulated by Directives 2001/18/EC and 2009/41/EC of the European Parliament and of the Council and their national implementation in the member States respectively. This legislation obliges Merck to request certain information about you and the establishment where the GMOs are being handled. Click here for Enduser Declaration (EUD) Form. Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Product Usage Statements

Quality Assurance

Evaluated by transduction of HT-1080 cells and fluorescent imaging performed for assessment of target localization and transduction efficiency.

Usage Statement

This product contains genetically modified organisms (GMO). Within the EU GMOs are regulated by Directives 2001/18/EC and 2009/41/EC of the European Parliament and of the Council and their national implementation in the member States respectively. This legislation obliges Merck to request certain information about you and the establishment where the GMOs are being handled. Click here for Enduser Declaration (EUD) Form.
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Storage and Shipping Information
Storage Conditions	Storage and Handling Lentivirus is stable for at least 4 months from date of receipt when stored at -80°C. After first thaw, place immediately on ice and freeze in working aliquots at -80°C. Frozen aliquots may be stored for at least 2 months. Further freeze/thaws may result in decreased virus titer and transduction efficiency. IMPORTANT SAFETY NOTE Replication-defective lentiviral vectors, such as the 3rd Generation vector provided in this product, are not known to cause any diseases in humans or animals. However, lentiviruses can integrate into the host cell genome and thus pose some risk of insertional mutagenesis. Material is a Risk Group 2 and should be handled under BSL2 controls. A detailed discussion of biosafety of lentiviral vectors is provided in Pauwels, K. et al. (2009). State-of-the-art lentiviral vectors for research use: Risk assessment and biosafety recommendations. Curr. Gene Ther. 9: 459-474.

Storage and Shipping Information

Storage Conditions

Storage and Handling
Lentivirus is stable for at least 4 months from date of receipt when stored at -80°C. After first thaw, place immediately on ice and freeze in working aliquots at -80°C. Frozen aliquots may be stored for at least 2 months. Further freeze/thaws may result in decreased virus titer and transduction efficiency.

IMPORTANT SAFETY NOTE
Replication-defective lentiviral vectors, such as the 3rd Generation vector provided in this product, are not known to cause any diseases in humans or animals. However, lentiviruses can integrate into the host cell genome and thus pose some risk of insertional mutagenesis. Material is a Risk Group 2 and should be handled under BSL2 controls. A detailed discussion of biosafety of lentiviral vectors is provided in Pauwels, K. et al. (2009). State-of-the-art lentiviral vectors for research use: Risk assessment and biosafety recommendations. Curr. Gene Ther. 9: 459-474.

Packaging Information
Material Size	1 vial (minimum of 3 x 10E8 IFU/mL)

Transport Information

Supplemental Information

Specifications

Global Trade Item Number
Bestellnummer	GTIN
17-10228	04053252595936

Documentation

LentiBrite™ Histone H2B-RFP Lentiviral Biosensor SDB

Titel
Sicherheitsdatenblatt (SDB)

LentiBrite™ Histone H2B-RFP Lentiviral Biosensor Analysenzertifikate

Titel	Chargennummer
LentiBrite Histone H2B-RFP Lentiviral Biosensor - Q1995263	Q1995263
LentiBrite Histone H2B-RFP -2662280	2662280
LentiBrite Histone H2B-RFP Lentiviral Biosensor	2438642
LentiBrite Histone H2B-RFP Lentiviral Biosensor - 3466668	3466668
LentiBrite Histone H2B-RFP Lentiviral Biosensor - 3520990	3520990
LentiBrite Histone H2B-RFP Lentiviral Biosensor - 3726107	3726107
LentiBrite Histone H2B-RFP Lentiviral Biosensor - 3886015	3886015
LentiBrite Histone H2B-RFP Lentiviral Biosensor - 3932364	3932364
LentiBrite Histone H2B-RFP Lentiviral Biosensor - 4076704	4076704
LentiBrite Histone H2B-RFP Lentiviral Biosensor - 4113827	4113827

Broschüre

Titel
New Products: Volume 3, 2012

Verwandte Produkte nach: Brand Facete

Kategorien

Life Science Research > Cell Analysis > Cell-based Assays > Lentiviral Biosensors

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