in-vivo bioassay

Hi everybody !!!!!!!!!
This topic very interesting by performing based.
Also will be useful for scientist working with recombinant protein. This is a pharmacodynamics type of assay where we have to measure any dynamic activity of animals.
With this introduction I am submitting my problem.
I am trying to validation of bioassay in female balb/c mice, what I said in above. I am doing on assay with single injection of standard solution EPO, remain things are similar to Ph EU. monogram, but I am found large variation of retic count in inter group, so I am always failing in fudicial limit while calculating assay. As per Ph. Eu. other parameter like regression, linearity and parallelism are found ok but regression value found very low that is 7-15 only.
So, My question are
1) Is animal play role for failing of fudicial limit? how and why?
2) How I control fudicial limit?

Please try to guide me in these era
please join in this discussion
Ramesh

Is the size (weight) of the mice accounted for? Is there an age range or sex of the mice specified for the assay? Among these variables, I think the size of the mouse is most important. Older male mice can vary a lot in size within a cage and that could affect the variance in your data.

We have try to manage/control age and weight also. 6-8 week old and 2 gms SD for weight. Although we are getting such variations

Hi Doctor,
Daily treatment of rats with 2,4-dithiobiuret (DTB, 1 mg/kg) produces an apparent flaccid muscle weakness that begins in the hindlimbs and ascends to involve muscles of the trunk, forelimbs, head, and neck. The insidious onset and progressive nature of the apparent weakness were detected by impaired rotarod performance, loss of righting response, and death after median cumulative doses of 4.9, 7.1, and 8.0 mg/kg, respectively, in male rats. If DTB treatment was stopped after the first day of rotarod failure, male rats recovered from impaired rotarod performance in 2–5 days. Sensitivity to DTB-induced rotarod failure was greatest in older rats of both sexes. Females were more susceptible than males to DTB-induced rotarod failure and frequently died after treatment was stopped, whereas males recovered. Effects of chronic DTB treatment that accompanied the impairment in rotarod performance, but did not appear to cause it, were decreased food and water intake, loss of body weight, diuresis, chromodacryorrhea, and watery, mucoid feces. Whole blood cholinesterase activity was not significantly altered and tail flick, corneal, vibrissae, pinna, and foot with-drawal responses could be elicited in animals that failed the rotarod test. The LD50 determined 48 hr after a single dose of DTB was 29 mg/kg yet acute treatment with up to 20 mg/kg did not affect rotarod performance when rats were tested for up to 3 weeks after treatment. The site and mechanism by which DTB impairs rotarod performance in the rat is unknown.

Three strains of the Indian Meal Moth,Plodia interpunctella, were compared in terms of their response to a granulosis virus under different environmental conditions. A significant difference in the relative susceptibilities to the virus of the three laboratory strains was established. Evidence of potential trade-offs with resistance was found in overall fecundity, pupal size and mortality at adult emergence. There was however little evidence that a reduction in resource level led to more trade-offs being apparent. No clear relationship was found between resistance to the lethal effects of the virus and susceptibility to the sublethal effects of the virus

Varition is common that is dependent on body weight. most important thing is mice size (weight),age and sex are accounted for assay.
Tirumal

Dear frnds,

I am looking forward to validate an in-vivo bioassay for EPO in normocythaemic mice, i would be greatful to have some literatures regarding the same.

Thanks and regards

Jith