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In vivo delivery protocol

FANA antisense oligonucleotides (ASOs) for RNA silencing and regulation

In vivo delivery protocol

The following protocol can be used for AUMsilence, AUMantagomir, AUMmirblocker, AUMmimic, AUMlnc, AUMskip and AUMblock.

Lipid-based transfection and electroporation are widely utilized, conventional methods to deliver siRNA into the cells. However, in many primary cells, particularly immune cells, hematopoietic cells and neurons, lipid reagents and electroporation are associated with high toxicity and poor transfection efficiency. Alternative delivery methods, such as viral vectors, require laborious optimization and viral production steps, and carry associated risk of genome integration.

The in vivo-ready FANA Antisense Oligonucleotides (FANA ASOs) are highly purified for direct delivery into animal models. FANA ASOs have unique, state-of-the-art chemical modifications that allow for simple and highly efficient delivery into animals without the need of special formulations. FANA’s modifications allow them to be efficiently taken up by the target tissues, and significantly increase their stability and their resistance to endonucleases.

FANA Delivery in Mice: Protocol

Note: The following is a general protocol for the use of FANA in mouse models. The protocol can be adapted for use in other mammalian or in vivo models.

  • 1. Prepare a FANA stock solution by reconstituting lyophilized ASOs at the desired concentration.

    • Resuspend FANA ASOs using the appropriate volume of sterile water or saline buffer.

    • We recommend a stock solution of 5-10 mg/ml. The approximate molecular weight of a 21 nt singlestranded FANA oligo is 6000-7000 g/mol (sequence-dependent). 100 nmol of FANA oligos is equivalent to 650 µg.

    • Pipette solution up and down 3-5 times while avoiding introduction of bubbles. Let the vial sit at room temperature for 5-10 min. Briefly centrifuge to collect solution to bottom of the tube.

  • 2. Calculate FANA concentrations for in vivo delivery.

    • For in vivo systemic delivery, we recommend a dose between 3 – 30 mg/kg in mice. If topical (organ, intratumoral, etc) delivery is desired, the concentration can be adjusted accordingly, considering the volume of the organ or tumor.

    • We recommend you perform a dose response using 2-3 working concentrations to determine the most optimum concentration for your specific application.

    • If long-term gene silencing is desired, multiple doses of FANA can be administered.

  • 3. Administer FANA oligos to mice.

    • FANA oligos can be administered via different routes: intravenous (IV), intraperitoneal (IP), intradermal (ID), intratumoral (IT), intranasal (IN), intratracheal, hydrodynamic tail injection, inhalation, or local organ delivery.

  • 4. Assay for gene knockdown in your target tissues.

    • Note: Uptake of fluorescently-labeled FANA can be observed as early as 24 hr, but full knockdown in vivo is best assessed at 72 hr to several days post FANA treatment.

FANA In Vivo Delivery Protocol at a Glance

Additional Notes:
  1. Depending upon the experiment different time points can be used to measure knock down or related effects for up to several days using a single dose (and weeks in some cases).

  2. If prolonged knockdown is desired, FANA oligos can be re-administered every few days.

  3. FANA oligos can be fluorescently labeled (or with any desired label) to monitor cellular uptake or biodistribution.

  1. FANA ASOs are shipped in lyophilized form. Upon arrival, store them in -20°C.

  2. When ready to use, resuspend FANA ASOs in sterile water or your favorite buffer at the desired concentration. Aliquot resuspended FANAs in working aliquots to avoid multiple freeze-thaw cycles.

Contact and customer support: For questions on products, protocols or new orders please reach out to us or visit us at .

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