PT-67

The PT-67 cell line is a murine retrovirus packaging cell line commonly used for the production of retroviral vectors for gene delivery purposes. Derived from NIH 3T3 cells, PT-67 expresses viral structural proteins and envelope proteins necessary for the assembly of retroviral particles. Notably, it is optimized to produce vectors pseudotyped with the 10A1 murine leukemia virus (MLV) envelope protein, which enables entry into cells using multiple phosphate transport protein receptors, including the gibbon ape leukemia virus receptor (Glvr-1 or Pit-1) and the amphotropic receptor Ram-1 (Pit-2).

The unique feature of PT-67 packaging cells lies in the 10A1 envelope protein's ability to utilize both Glvr-1 and Ram-1 receptors. This dual receptor specificity significantly expands the host range of retroviral vectors produced by PT-67, making it advantageous for transducing a variety of cell types across species, including human, mouse, and rat cells. For instance, the 10A1-pseudotyped retroviral vectors can efficiently transduce murine NIH 3T3 cells and human fibroblasts by leveraging these receptors. However, receptor usage patterns are species- and tissue-specific, as observed in studies where 10A1 pseudotyped vectors efficiently use Ram-1 but show variable efficiency with Glvr-1 in different human and rodent cell lines.

Retroviral vectors produced using PT-67 cells are helper virus-free, ensuring safety and efficiency in gene transfer experiments. This makes PT-67 a valuable tool for stable gene delivery in experimental and therapeutic applications, particularly where broad host range transduction is desired. The development and optimization of PT-67 packaging cells have facilitated advancements in gene therapy, enabling the delivery of therapeutic genes into target cells with enhanced specificity and efficiency.

37°C, 5% CO₂, humidified atmosphere.

1. Confirm that the vial remains deeply frozen upon delivery, as cells are shipped on dry ice to maintain optimal temperatures during transit.

2. Upon receipt, either store the cryovial immediately at temperatures below -150°C to ensure the preservation of cellular integrity, or proceed to step 3 if immediate culturing is required.

3. For immediate culturing, swiftly thaw the vial by immersing it in a 37°C water bath with clean water and an antimicrobial agent, agitating gently for 40-60 seconds until a small ice clump remains.

4. Perform all subsequent steps under sterile conditions in a flow hood, disinfecting the cryovial with 70% ethanol before opening.

5. Carefully open the disinfected vial and transfer the cell suspension into a 15 ml centrifuge tube containing 8 ml of room-temperature culture medium, mixing gently.

6. Centrifuge the mixture at 300 x g for 3 minutes to separate the cells and carefully discard the supernatant containing residual freezing medium.

7. Gently resuspend the cell pellet in 10 ml of fresh culture medium. For adherent cells, divide the suspension between two T25 culture flasks; for suspension cultures, transfer all the medium into one T25 flask to promote effective cell interaction and growth.

8. Adhere to established subculture protocols for continued growth and maintenance of the cell line, ensuring reliable experimental outcomes.

Mycoplasma contamination is excluded using both PCR-based assays and luminescence-based mycoplasma detection methods.

To ensure there is no bacterial, fungal, or yeast contamination, cell cultures are subjected to daily visual inspections.

For long-term preservation, place vials in vapor-phase liquid nitrogen at about −150 to −196 °C. Storage at −80 °C is acceptable only as a short interim step before transfer to liquid nitrogen.

Cryopreserved cell lines are shipped on dry ice in validated, insulated packaging with sufficient refrigerant to maintain approximately −78 °C throughout transit. On receipt, inspect the container immediately and transfer vials without delay to appropriate storage.

At Cytion, we stand behind the quality and viability of our products. We guarantee that our cells will be viable at the start of culture for 60 days after shipment from our facilities. We assure that all materials conform to the specifications described in the Certificate of Analysis and/or Product Sheet.

For full details, please refer to our Warranty Statement.