Dear Editor,

We read with interest the paper published by Boschi, et. al.[¹] in which immunohistochemistry was performed on orbital tissues from patients with thyroid associated ophthalmopathy (TAO) and compared to non-diseased orbital tissue.

Our laboratory recently reported positive TSH receptor staining within normal human muscle fibres, using one of the same antibodies (3G4) as Boschi et. al., (supplied by Costagliola) and a commercial antibody (3B12) [²].

Our findings differ from Boschi et. al.’s as no staining of the muscle fibres was visible in their experience.

Assessing the techniques used gave some possibilities as to why our findings differ. Our paraffin-embedded tissues were subjected to a proteolytic antigen retrieval step, as commonly used in avidin-biotin staining.[³] The reason for this is that formalin used in fixation is notorious for altering protein immunoreactivity, and hence masking protein expression.[⁴][⁵]

Moreover, the amplification immunohistochemistry kit used in our experiments is possibly more sensitive than conventional immunohistochemistry used in the experiments of Boschi, et. al.[⁶]

We do not dispute the finding that TSHR expression is elevated in orbital connective tissue of diseased patients. Combined with our findings, Boschi et. al.’s paper also suggests that expression of TSH-R on normal muscle fibres is lower than in the connective tissue of diseased patients. Boschi et. al. has successfully produced more evidence that connective tissues in the orbit are active in TAO affected patients, however the potential role of the extraocular muscle in the pathogenesis of TAO should also be considered.

Yours sincerely,

Steven J. Kloprogge and Albert G. Frauman

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