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We have generated an induced KO mice line for a specific gene. However when i perform IF staining I can still see substantial levels of the protein for this specific gene produced, even though the Genotyping results indicate that the gene KO was successfully performed. Did anyone else have the same problem? or can you think of a plausible explanation to this problem? Thank you in advance!

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    $\begingroup$ How is the knockout induced? When was it induced? When are you measuring protein? $\endgroup$
    – Bryan Krause
    Apr 12 at 14:19
  • $\begingroup$ Mice are induced by a 16 day administration of doxycycline via the drinking water. The study model is then initiated after a 7-days wash-out (restore to normal water) phase. I measure the protein at day 10, so in total 17 days after the interruption of induction. $\endgroup$
    – shaii
    Apr 12 at 14:28
  • $\begingroup$ How are you measuring protein levels? What controls have you performed? $\endgroup$
    – Bryan Krause
    Apr 12 at 14:29
  • $\begingroup$ At the moment, running the initial phase of the experiments, we are comparing the protein expression to the wild type animals by performing the same IF staining on both groups. The protein signal is not significantly different in KO mice as compared to the wild type ones. $\endgroup$
    – shaii
    Apr 12 at 14:32
  • $\begingroup$ Are the knockouts homozygous? Is the antibody known to be specific enough for this particular protein, without affinity to related proteins? Do you know anything about the half-life of the protein? $\endgroup$
    – Bryan Krause
    Apr 12 at 14:35

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