It is possible that this is due to "Biotin nuclei" where excess biotin is found in the nuclei of some cells, see below:
Optically clear nuclei have been reported in endometrial epithelium associated with first and second trimester abortions (Sickel & di Sant'Agnese 1994). Optically clear nuclei have also been found in different types of tissues of diverse organs such as ovary, thyroid and lung (Nakatani et al 1994, Mount & Cooper 2001). The optically clear nuclei contain excess biotin. Endogenous biotin immunoreactivity is generally not visualized in formalin fixed, paraffin-embedded tissues unless a heat-induced antigen retrieval step has been introduced (Mount & Cooper 2001). In this placental section, optically clear nuclei (containing biotin) bind to the streptavidin of the ABC technique giving a reaction similar to that seen with CMV containing cells. If a polymer method (or even the original Sternberger's PAP method) is used then this anomalous staining will disappear, thus allowing confident demonstration of CMV infected nuclei. The false-positive staining pattern caused by endogenous biotin can be cytoplasmic or nuclear. A report of positive immunoreactivity of hepatocellular carcinomas for inhibin was later determined to be a false-positive finding due to cytoplasmic endogenous biotin. Steroid cell tumours of the ovary were found to demonstrate endogenous biotin cytoplasmic staining in 36% of cases. Immunoreactivity for anti-Herpes virus immunohistochemical staining in a series of endometria was also later determined to be a false-positive result due to biotin. The prominent intranuclear inclusions, resembling herpes virus cytopathic effect, were caused by intranuclear biotin and not viral particles. Similar false positive staining for CMV in products of conception has also been reported (Mount & Cooper 2001). False-positive staining can be cytoplasmic or nuclear. When cytoplasmic, the appearance of the false signal is that of a dull brown granular or fluffy staining pattern. If this quality of staining is observed with several different antibodies, endogenous staining by biotin should be considered. When nuclear, a false-positive reaction may be associated with optically clear nuclei identified on H&E stained sections. False-positive staining due to endogenous biotin, however, does not occur in a cell membrane pattern (Mount & Cooper 2001). Mount SL & Cooper K (2001) "Beware of biotin: a source of false-positive immunohistochemistry" Current Diagnostic Pathology 7:161-167. Nakatani et al (1994) Am J Surg Pathol 18(6):637-642. Sickel & di Sant'Agnese (1994) Arch Pathol Lab Med 118:831-833 Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA) Laboratory Manager & Senior Scientist Tel: 612 9845 3306 Fax: 612 9845 3318 the children's hospital at westmead Cnr Hawkesbury Road and Hainsworth Street, Westmead Locked Bag 4001, Westmead NSW 2145, AUSTRALIA -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Eva Permaul Sent: Monday, 23 July 2012 11:40 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Secondary antibody causing nuclear staining? Hello, I have noticed that our biotinylated secondary antibodies on occasion cause nuclear staining in some samples. Why is this? It is not every time so I find it rather stange. Anyone know why this is happening and what I can do to avoid it? Thank you for any suggestion, Eva Permaul Georgetown University _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ********************************************************************************* This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, The Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. ********************************************************************************* _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
