Lentivirus LV User Manual

Table of Contents

Delivery                         02

Shipping                        02

Storage and Handling            02

Safety Information               02

In Vitro Applications             03

Adherent Cell Infection         04

Suspension Cell Infection         05

Note: First-time users must read this manual carefully before proceeding with any operations!

Delivery

Shipping:

The delivered LV virus is stored in PBS buffer and shipped on dry ice. Upon receipt, if there are any issues such as damaged packaging, please contact our staff promptly.

Storage and Handling:

1. Upon receipt, if only a small volume will be used, aliquot the virus into smaller volumes (use DNase/RNase-Free sterile consumables and seal with parafilm). Store at -80°C for up to approximately one year; at -20°C for 2–3 weeks; at 4°C for 1–2 weeks. After one year of storage, please re-titer the virus before use.

  2. The virus can be diluted with sterile PBS or sterile saline. Please prepare the working dilution freshly before use according to the actual required volume.

  3. Thaw the virus on ice before use and keep it on ice throughout the experiment. If the unused virus is stored at 4°C, please use it within one week.

Note: Avoid repeated freeze-thaw cycles, as this will significantly reduce the viral titer.

Safety Information:

1. Before using the virus, operators should be fully aware of the safety issues associated with virus-related experiments and enhance safety awareness. During experiments, please be sure to wear lab coats, masks, goggles, and disposable gloves. 

2. It is recommended to perform virus operations in a Class II biological safety cabinet according to BSL-2 (Biosafety Level-2) guidelines. (If using a clean bench, do not turn on the exhaust fan to avoid contact between the virus and the external environment.) 

3. During virus handling, avoid protease or bacterial contamination. Use DNase/RNase-Free consumables for all operations. 

4. If eyes, skin, etc. accidentally come into contact with the virus, rinse immediately with plenty of water. If the virus contacts wounds or other sites, immediately scrub with 10% povidone-iodine solution, then rinse thoroughly with plenty of water for at least 20 minutes. 

5. After the experiment, inactivate the virus with freshly prepared 1% sodium hypochlorite solution for at least 20 minutes. Alternatively, 2% glutaraldehyde solution or 0.25% SDS solution can be used, or autoclaving (121°C, 1 hour) may be employed. 

6. After the experiment, wash hands thoroughly with soap and water. All biological waste disposal must comply with regulations issued by public agencies and the specific institution.

In Vitro Applications:

Viral titer refers to the number of active viral units per unit volume of virus solution and is a key indicator of virus quality. The titer unit for LV is TU (transducing units)/mL. Multiplicity of infection (MOI) is the ratio of virus to cells during infection, i.e., MOI = (virus titer × virus volume) / cell number. The optimal MOI for a given cell type is the MOI at which approximately 80% of the cells are infected. For LV, the MOI unit is TU/cell.

For first-time lentivirus users, it is recommended to perform a gradient pre-experiment to determine the optimal infection conditions (e.g., optimal MOI) for your target cells.

Lentivirus Protocol:

1. Cell Seeding: According to the recommended seeding volumes in the table below, count healthy cells and seed the appropriate number into the corresponding culture vessels. Incubate at 37°C for 16–24 hours until cells reach 60–80% confluence.

2. Virus Infection: Based on pre-experiment results, select the appropriate MOI and infection conditions. Add the corresponding amount of virus to the cell culture system (to increase virus-cell contact, the culture volume may be reduced appropriately). Then place the cells back into the incubator for infection.

3. Medium Change: After 12–20 hours, check cell morphology and replace with fresh medium as needed (it is recommended to check cell status after 8 hours; if cells are unhealthy, medium change may be performed earlier to maintain normal growth). Do not exceed 24 hours before changing medium.

4. Evaluate Infection Efficiency: 48–72 hours post-infection, observe fluorescent protein expression under a fluorescence microscope to assess infection efficiency.