Giving her verdict, Judge Claire Sharp said John Atkinson did not lose his job because of his disability.
Mr Atkinson was dismissed, as were all of Mr Gill's staff, in April last year.
Some of the employees were later re-hired and Mr Atkinson had argued he was not given a job because he needed a hip operation.
But the claim at a tribunal in Cardiff was thrown out.
Ms Sharp said: "The reason that the claimant lost his job had nothing to do with his disability".
"It was to do with the political situation and Mr Gill's view that it would be better to be the only MEP without any staff," she told the hearing.
The tribunal had earlier heard that Mr Gill was concerned in the run up to the 2016 assembly election about the possibility that Olaf - an EU fraud body - would investigate him and his staff.
The MEP wanted to avoid "cross over" between political campaigning and EU funding.
Mr Gill, who was elected to the assembly and now sits as an independent AM, had wanted to stand down as an MEP but said there were a "huge catalogue" of reasons why he changed his mind.
Mr Atkinson - a 67-year-old former Royal Marine from Llanpumsaint, Carmarthenshire - was a campaign manager for Mr Gill, before becoming a constituency manager after he got elected in 2014.
In response to the ruling, Mr Gill said: "Employment tribunals serve a very important function in protecting workers from bad employers. I was never, and have never been, a bad employer."
A former employee of Nathan Gill has lost a claim of disability discrimination against the UKIP Wales MEP at an employment tribunal.