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Nuclear localization of lactoferrin in the human granulocyte: Artifact incurred during slide preparation
Abstract:
Within the blood cells, lactoferrin is found only in the late stage neutrophilic granulocytes. Lactoferrin first appears in these cells during the myelocyte stage of development coincidentally with the specific or secondary granules. Most investigators report a cytoplasmic immunocytochemical localization reaction within the granulocyte. However, others have observed a prominent nuclear localization reaction. Treating the cells with certain fixatives was shown to prevent the relocation of lactoferrin from the cytoplasm to the nucleus when the localization was done on granulocytes prepared by smearing. The present study demonstrated that the relocation of lactoferrin is only a problem when cells were smeared or cytocentrifuged onto slides or fractionated for the purpose of isolating cellular organelles. Under these conditions the selection of fixative is an important consideration. Exposing isolated lactoferrin to a fixative effective in retaining lactoferrin in the cytoplasm of granulocytes smeared on slides did not alter a number of its physical properties. The results suggest that maintenance of the normal cytoarchitecture or effect of fixative on other cellular components prevents the relocation of lactoferrin within the cell during tissue processing and the direct action of fixation on lactoferrin is probably not responsible for this effect.
Insights
Lactoferrin relocation within neutrophils during sample preparation is an artifact. Proper tissue processing and fixation prevent this nuclear shift, preserving its true cytoplasmic localization.
Area of Science:
- Hematology
- Cell Biology
- Immunocytochemistry
Background:
- Lactoferrin is a key protein found in late-stage neutrophilic granulocytes.
- Its localization within these cells is debated, with reports of both cytoplasmic and nuclear presence.
- Previous studies suggest fixatives influence lactoferrin localization in smeared granulocytes.
Purpose of the Study:
- To investigate the cause of lactoferrin relocation within neutrophils during sample preparation.
- To determine if fixation directly causes lactoferrin's nuclear translocation.
- To identify optimal methods for preserving lactoferrin's native cellular location.
Main Methods:
- Examining lactoferrin localization in granulocytes subjected to smearing, cytocentrifugation, and organelle isolation.
- Testing the effect of specific fixatives on lactoferrin localization in processed cells.
- Assessing the impact of fixation on the physical properties of isolated lactoferrin.
Main Results:
- Lactoferrin relocation to the nucleus is an artifact primarily occurring during smearing, cytocentrifugation, or fractionation.
- The choice of fixative is critical when these processing methods are used.
- Fixation did not alter the physical properties of isolated lactoferrin, suggesting fixation itself is not the direct cause of relocation.
Conclusions:
- The observed nuclear localization of lactoferrin is an artifact of sample preparation, not a native cellular event.
- Maintaining normal cytoarchitecture or interactions with other cellular components prevents relocation.
- Fixation's direct effect on lactoferrin is unlikely to be the cause of its translocation.