LAG3 CRISPR/Cas9 Lentivirus (Non-Integrating)

Catalog #

78060

$995 *

Size: 500 µl x 2

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Description

Lymphocyte-activation gene 3 (LAG3, CD223) is a cell surface protein that belongs to the immunoglobulin (Ig) superfamily. LAG3 is expressed on activated T-cells, Natural Killer cells, B-cells, and plasmacytoid dendritic cells. Its main ligand is the MHC class II, to which it binds with higher affinity than CD4. It negatively regulates cellular proliferation, activation, and homeostasis of T-cells in a similar fashion as CTLA-4 and PD-1, and has been reported to play a role in T-reg suppressive function. A number of LAG3 antibodies are in preclinical development for the treatment of cancer and autoimmune disorders. LAG3 may be a better immune checkpoint inhibitor target than CTLA-4 or PD-1, because antibodies targeting CTLA-4 or PD-1 only activate effector T-cells while failing to inhibit T-reg activity, whereas an antagonist LAG3 antibody can both activate effector T-cells (by downregulating the LAG3 inhibiting signal) and inhibit induced (i.e. antigen-specific) T-reg suppressive activity.

The LAG3 CRISPR Lentiviruses are replication incompetent, HIV-based, VSV-G pseudo-typed lentiviral particles that are ready to be transduced into almost all types of mammalian cells, including primary and non-dividing cells. The particles contain a CRISPR/Cas9 gene driven by an EF1A promoter, along with 4 sgRNA (single guide RNA) targeting human LAG3 (GenBank Accession #NM_002286) driven by a U6 promoter (Figures 1 and 2).

Schematic of the Lenti-vector used to generate the LAG3 CRISPR/Cas9 Lentivirus.

Figure 1. Schematic of the Lenti-vector used to generate the LAG3 CRISPR/Cas9 Lentivirus.

Gene Target:	Primer ID:	sgRNA Sequence
LAG3	LAG3-1	GTTCCGGAACCAATGCACAG
LAG3	LAG3-2	GGGAGTTACCCAGAACAGTG
LAG3	LAG3-3	CGTCCCGCCCCACATACTCG
LAG3	LAG3-4	GCTCACATCCTCTAGTCGAA

Figure 2. List of sgRNA Sequences in the LAG3 CRISPR/Cas9 Lentivirus.

Note: unlike Human LAG3 CRISPR/Cas9 Lentivirus (Integrating) (BPS Bioscience, #78053), the Human LAG3 CRISPR/Cas9 Lentivirus (Non-Integrating) is made with a mutated Integrase, resulting in only transient expression of the Cas9 and LAG3-targeting sgRNA. While this may minimize potential off-targeting risks due to either prolonged expression or integration of the Cas9, puromycin selection should not be used for more than 48 hours post-transduction, which may lower knockout efficiency.

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Synonyms

Lymphocyte-activation gene 3 CRISPR, CD223 Lentivirus

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Product Info

Storage and Usage

Citations

Supplied As

Two vials (500 μl x 2) of lentivirus at a titer ≥1 x 10⁶ TU/ml. The titer will vary with each lot; the exact value is provided with each shipment.

Formulation

The lentiviruses were produced from HEK293T cells in medium containing 90% DMEM + 10% FBS.

Storage/Stability

Lentiviruses are shipped with dry ice. For long-term storage, it is recommended to store the lentiviruses at -80°C for up to 12 months from date of receipt. Avoid repeated freeze/thaw cycles. Titers can drop significantly with each freeze/thaw cycle.

Instructions for Use

See assay protocol for detailed instructions.

Applications

1. Transient knock-down of LAG3 in a target cell pool.
2. Generation of stable LAG3 knock-out cells using transient puromycin selection (48h maximum) followed by limited dilution.

Shipping Temperature

-80°C

Notes

The CRISPR/CAS9 technology is covered under numerous patents, including U.S. Patent Nos. 8,697,359 and 8,771,945, as well as corresponding foreign patents applications, and patent rights.

Warnings

Biosafety
None of the HIV genes (gag, pol, rev) will be expressed in the transduced cells. Although the pseudotyped lentiviruses are replication-incompetent, they do require the use of a Biosafety Level 2 facility. BPS recommends following all federal, state, local, and institutional regulations and using all appropriate safety precautions.

Scientific Category

Immunotherapy

Regulatory Status

Research Use Only

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LAG3 CRISPR/Cas9 Lentivirus (Non-Integrating)

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