Intravitreal Infliximab Clearance in a Rabbit Model: different sampling methods and assay techniques
Fabrizio Giansanti 1*,
Matteo Ramazzotti 2,
Matteo Giuntoli 3,
Gianni Virgili 4,
Lorenzo Vannozzi 3,
Donatella Degl'Innocenti 2,
and
ugo menchini 5
1 Department of oto-neuro-Ophthalmological Surgical Sciences, University of Florence, V.le Morgagni 85, Florence, 50134, Italy
2 Department of Biochemical Sciences, University of Florence, Florence, Italy
3 Department of oto-neuro-Ophthalmological Surgical Sciences, University of Florence, Florence, Italy
4 Ophthalmology, University of Florence, Florence, Italy; Department of oto-neuro-Ophthalmological Surgical Sciences, University of Florence, Florence, Italy
5 Department of oto-neuro-Ophthalmological Surgical Sciences, University of Florence, Florence, Florence, Italy
* To whom correspondence should be addressed. E-mail: fabriziogiansanti{at}interfree.it.
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Abstract |
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Purpose. To investigate the clearance of intravitreal infliximab using different sampling techniques and immunoassay methods in rabbits.
Methods. 1.6 mg of infliximab was intravitreally injected into both eyes of 47 rabbits. Two approaches were used to collect the vitreous; the classic method and a micro-sampling technique. Whereas the classic method consists in the whole vitreous being collected after enucleation, the micro-sampling technique consisted in the aspiration of small samples of 10-15 µl with a 200 µl syringe. Samples were taken from 30 min to 40 days using both methods, which were then compared. Infliximab concentration was estimated using competitive ELISA, dot-blot and Western blot.
Results. The vitreous half-life of infliximab was estimated to be 6.5 ± 0.6 days. Our data indicated a mono-exponential decay, reaching its conclusion after about 40 days. This decay was preceded by a four-day long diffusion in the vitreous. Micro-sampling proved to be effective in the vitreous collection, giving statistically comparable signals (± 4%, p=0.68) with respect to the classic procedure. ELISA proved to be the best analytical technique - especially if coupled with micro-samplings - because of its lower detection limit, precision and the reduced amount of sample needed. No differences were observed between half-life values obtained by ELISA and dot-blot (p=0.081) and Western blot (p=0.614).
Conclusion. This study improved our knowledge of infliximab clearance in the vitreous and confirmed the validity of a micro-sampling technique that was compared with the classic one. ELISA was found to be the best analytical technique when using micro-sampling.
Key Words:
pharmacokinetics, intravitreal drug delivery, uveitis
