Background: Antibody validation for tissue staining is required for reproducibility; criteria to ensure validity have been published recently. The majority of these recommendations imply the use of routinely processed (formalin-fixed, paraffin-embedded) tissue. Materials & methods: We applied to lightly fixed frozen sections a panel of 126 antibodies validated for formalin-fixed, paraffin-embedded tissue with extended criteria. Results: Less than 30% of the antibodies performed as expected with all fixations. 35% preferred one fixation over another, 13% gave nonspecific staining and 23% did not stain at all. Conclusion: Individual antibody variability of the paratope’s fitness for the fixed antigen may be the cause. Revalidation of established antibody panels is required when they are applied to sections whose fixation and processing are different from the tissue where they were initially validated.

Bolognesi, M., Mascadri, F., Furia, L., Faretta, M., Bosisio, F., Cattoretti, G. (2021). Antibodies validated for routinely processed tissues stain frozen sections unpredictably. BIOTECHNIQUES, 70(3), 137-148 [10.2144/btn-2020-0149].

Antibodies validated for routinely processed tissues stain frozen sections unpredictably

Bolognesi, Maddalena M
Primo
;
Bosisio, Francesca M;Cattoretti, Giorgio
Ultimo
2021

Abstract

Background: Antibody validation for tissue staining is required for reproducibility; criteria to ensure validity have been published recently. The majority of these recommendations imply the use of routinely processed (formalin-fixed, paraffin-embedded) tissue. Materials & methods: We applied to lightly fixed frozen sections a panel of 126 antibodies validated for formalin-fixed, paraffin-embedded tissue with extended criteria. Results: Less than 30% of the antibodies performed as expected with all fixations. 35% preferred one fixation over another, 13% gave nonspecific staining and 23% did not stain at all. Conclusion: Individual antibody variability of the paratope’s fitness for the fixed antigen may be the cause. Revalidation of established antibody panels is required when they are applied to sections whose fixation and processing are different from the tissue where they were initially validated.
Articolo in rivista - Articolo scientifico
Antibody validation • antigen retrieval • epitope • fixatives • frozen sections • paratope;
mazze di tamburo;
English
5-feb-2021
2021
70
3
137
148
none
Bolognesi, M., Mascadri, F., Furia, L., Faretta, M., Bosisio, F., Cattoretti, G. (2021). Antibodies validated for routinely processed tissues stain frozen sections unpredictably. BIOTECHNIQUES, 70(3), 137-148 [10.2144/btn-2020-0149].
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/10281/301514
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