Papanicolaou Staining: A Technique to Visualize the Cytological Features of Tubal Cells
Published: April 30, 2023
Abstract
Source: Chen, H. et al. Tubal Cytology of the Fallopian Tube as a Promising Tool for Ovarian Cancer Early Detection. J. Vis. Exp. (2017).
In this video, we demonstrate a modified protocol of Papanicolaou staining to study tubal cytology. This technique can be used to visualize the cytological features of malignant and non-malignant fallopian tube epithelial cells.
Protocol
1. Modified Papanicolaou Staining Protocol
- Stain the tissue smeared slides by running the automatized non-gyn stain procedure, as illustrated in Table 1.
NOTE: The staining method represents a modification of the Papanicolaou staining technique that we use routinely for non-gynecologic specimens. We use an automated tissue processor to stain some slides. We chose the Non-Gyn staining procedure over the Gyn staining procedure in this study. The Eosin used is EA-65, instead of the EA-50 used for gynecological specimens (Pap smear specimen). In addition, OG-6 in the Gyn staining procedure, which is mainly for squamous cell staining, is not included in the staining procedure since no squamous cells are expected to be seen in this study. EA65 (Eosin Azure) is a counter stain solution comprised of 3 dyes: Eosin Y, Fast green FCF, and Bismarck brown Y. The procedure is illustrated in Table 1. - Perform a manual quick Pap stain procedure as follows:
- Dip the tissue smeared slide into 95% ethanol, 10 times, with each dip taking about 1 s.
- Dip the slide into H2O, 10 times, with each dip taking about 1 s.
- Emerge the slide in hematoxylin for 10 – 60 s.
- Dip the slide into H2O, 10 times, with each dip taking about 1 s.
- Dip the slide into 95% ethanol,10 times, with each dip taking about 1 s.
- Emerge the slide in EA65 for 1 – 3 min.
- Dip the slide into H2O, 10 times, with each dip taking about 1 s.
- Dip the slide into 95% ethanol, 10 times, with each dip taking about 1 s.
- Dip the slide in xylene until the slide becomes clear.
NOTE: This is a modified Papanicolaou staining procedure. This method is used as a more rapid stain method for cases with limited time or space like on-site fine needle aspiration and STAT specimens. This procedure provides comparable quality staining as the automatized Non-Gyn stain procedure. The procedure is illustrated in Table 2.
Table 1: Non-Gyn Stain Steps.
| 95% Ethanol | 5 min |
| H2O | 10 sec |
| Hematoxylin | 15 sec – 3 min |
| Distilled H2O | 5 min |
| 95% Ethanol | 30 sec |
| EA65 | 2 – 5 min |
| 95% Ethanol | 30 sec |
| 95% Ethanol | 30 sec |
| 100% Ethanol | 30 sec |
| 100% Ethanol | 30 sec |
| Xylene | 30 sec |
| Xylene | 5 min |
Table 2: Manual Quick Pap Stain Steps.
| 95% Ethanol | 10 dips |
| H2O | 10 dips |
| Hematoxylin | 10 sec-1 min |
| Distilled H2O | 10 dips |
| 95% Ethanol | 10 dips |
| EA65 | 1 – 3 min |
| 95% Ethanol | 10 dips |
| 100% Ethanol | 10 dips |
| Xylene | Dip until clear |
Divulgaciones
The authors have nothing to disclose.
Materials
| Papanicolaou Stain, EA 65 | sigma aldrich | HT40432 | |
| 95% Ethanol | sigma aldrich | 652261 | |
| 100% Ethanol | sigma aldrich | 493538 | |
| Xylene | sigma aldrich | 214736 | |
| Hematoxylin | sigma aldrich | H9627 |
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Citar este artículo
Papanicolaou Staining: A Technique to Visualize the Cytological Features of Tubal Cells. J. Vis. Exp. (Pending Publication), e21052, doi: (2023).