Custom Animal Ascites Production

Antagene offers ascites production from small Research & Development scale to large-scale production. We secured a USDA registered research facility, and we comply with all federal guidelines for care and use of laboratory animals.

Priming and cell preparation:
You may provide sufficient cells from designated cell line (approximately 2-2.5 x 106 cells per animal). A minimum of 5 mice will be used per hybridoma clone. As soon as we receive your order, we will start to prime at least 10 mice per hybridoma clone with Pristane for you. (Default: minimum 14 days of priming, with 0.5 ml of Pristane / mouse). The time interval between priming and inoculation of hybridoma cells is 10-14 days. Generally, very high concentrations of cells are associated with greater mortality and concentrations < 1 x 105 cells elicit fewer ascitic tumors and these tend to have a smaller volume yield. Cell suspensions should be prepared sterilely in physiological solutions.
If you prefer to send us frozen vials, 3 vials of cells (3×106 cells / vial) from designated cell line are required. As soon as we received the sample, Cells are thawed and viability is tested.  We then begin to expand cells to appropriate number for inoculation (Default: 2-3 X 106 cells/ animal). We determine percent viability via Trypan Blue Dye. Approximately one or two weeks are needed to prepare cells from frozen stocks for injection, and there is an additional charge ($300/clone) for this service.
(Additional: Nude and SCID mice are available for ascites production for non-murine hybridoma cell lines.)

Harvesting Ascites Fluid:
Hybridomas are injected into pristane-primed mice, then mice are monitored daily for ascites development and general condition. Ascites collection begins when accumulation of ascites becomes evident and continues until final harvests are performed. You can choose to receive unpurified, defibrinated ascites, or Antagene will purify ascites for you. In general, ascites production is completed within 2-3 weeks after injection of hybridomas.

1. Cells are harvested in logarithmic phase of growth and injected into mice.
2. Approximately 2 weeks after cell inoculation, tapping will begin. Mice will be held up to 6 weeks after cell injection.
3. Mice receive a maximum of 3 needle taps with a terminal tap.
4. Daily taps are pooled, and then centrifuged at low speed.
5. Daily taps are stored frozen at -70° C, in 50-ml centrifuge tubes.
6. At completion of tapping, centrifuge tubes from daily taps are thawed and pooled.
7. Final pooled material receives high-speed centrifugation.

Deliverable:
The ascites fluid will be sent back to you via overnight transporter. If you ask for antibody purification, we will use those ascites to purify antibody (additional charge $500 for IgG purification), 15-20mg purified antibodies will be delivered at the end.  Shipment costs to your laboratory are additional as indicated in our invoice.Confidentiality
We will keep no sample after the experiment achievement. No sample will be transferred to any third party or be commercially used. You will remain the only proprietary of the hybridoma. Moreover, we will not disclose to any third party that we have used your material for any manipulation or worked with your company. Starting with your own hybridoma, we cannot guarantee ascites quality in case of low antibody yield.

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Please contact us for a free consultation/ quotation of your customized Ascites Production.