Blue dye can be injected around the primary tumour or scar (in a similar way to how the radiocolloid was injected when adopting the peritumoural route) 10–20 min prior to the operation in a volume of 0.5–1 mL. The injection should be performed after the patient is anaesthetized to avoid a painful injection. Five minutes of gentle massage at the injection site enhances movement of the dye through the lymphatics to the SLN. Within 5–15 min, the SLN is coloured, with washout occurring after approximately 45 min. Currently, the most commonly used dyes are patent blue V, isosulfan blue and methylene blue.
FIG. 5.8. A 29 year old patient with inferior medial quadrant cancer in her left breast. Early images (anterior and oblique) show two separate lymphatic drainage pathways, respectively, to the axilla and to the internal mammary chain. When drainage to more than one anatomic region is seen, each of those regions must have at least one SLN.
Multiple studies have validated the use of blue dyes as markers for SLNs with high detection rates (ranging from 75% to 95%), although these are slightly lower than the rates achieved by radiocolloids. In most cases, the same SLNs are detected by blue dyes as are detected by radiocolloids. A notable disadvantage of using blue dyes instead of radiotracers is that blue dyes are not helpful if extra-axillary nodes (IMN or supraclavicular) are to be evaluated [5.80, 5.81].
There are some contraindications to the use of blue dyes. Blue dyes may interfere with pulse oximetry readings, so in certain patients, they should be used with caution. Furthermore, blue dye is contraindicated in pregnancy (because of the risk of an anaphylactic reaction), in patients with earlier allergic reactions to blue dye and in severe renal impairment (methylene blue) [5.82]. On the other hand, hypersensitivity reactions to radiocolloids are rare, but have also been reported.
Gamma detection probes must be able to detect the SLN within the exposed surgical cavity as well as from outside the skin surface. In fact, the first step of the procedure consists of confirming with external measurements the location of the SLN before making the surgical incision; this task requires the sensitivity of the detector to be sufficient to identify a weakly active SLN when attenuated by, typically, up to 5 cm of soft tissue. Discriminating activity counts within the SLN from those originating from nearby sites requires the probe to be well collimated with a small angle of view. Using the images and skin markings as guides, the probe (placed over the regions of highest counts) can be used to select the optimum location for incision. The probe is placed in a sterile
bag for intraoperative use in the surgical field. The surgeon uses the probe to guide dissection to the hot node(s) and places the probe in the surgical bed after node excision to confirm removal of the hot node(s). When working with the probe, it is important to direct the probe away from activity at the injection sites.
Counts are recorded per unit time with the probe in the surgical field, over the node before excision (in vivo) and after excision (ex vivo). A background tissue count is also recorded with the probe pointing away from the injection site, nodal activity or other physiological accumulation sites (i.e. liver) [5.42].
Deeply located SLNs are difficult to detect because of tissue attenuation;
furthermore, the large amount of radioactivity retained at the injection site may cause SLNs located nearby to be missed. This is frequently observed when the tumour is located in the upper outer quadrants, or in cases of IMN sentinel nodes when the tumour is located in the inner quadrants. In the latter instance, it is advisable to use thinner probes (e.g. 10 mm diameter) in the intercostal area to better depict the activity spot in an area with reduced surgical space.
5.4.3.1. Interpretation criteria
LNs of the first two categories defined in Section 4.5. (definitively or highly probable SLNs as identified during lymphoscintigraphy) must be removed for analysis. Low probability SLNs may sometimes be removed, depending on the amount of remaining radioactivity measured by the gamma probe. An SLN usually has at least ten times the background count, taken at a location remote from the injection site. Various probe criteria have been employed for identification of the SLN (e.g. counts per second recorded for the presumed SLN have been compared with non-SLNs in vivo, ex vivo or with background counts in vivo, considering as SLNs all those nodes with a count rate higher than 10% of the count rate of the hottest SLN).
When a hot SLN has been removed, the surgical bed should be checked for remaining activity. Even when one hot SLN is clearly noted on lymphoscintigraphy, this scintigraphic appearance could actually represent two close SLNs erroneously detected as one, owing to the limited spatial resolution of the gamma camera. LNs closer than approximately 15–20 mm may well appear as one single node, so in some cases, another hot node may still be present at a close location after removal of the hottest SLN. In this regard, the current use of SPECT/CT imaging is very helpful because it may provide information about the actual presence of a cluster of LNs rather than a single SLN. When other sources of activity are found in the lymphatic basin, the decision of whether to remove them will depend upon the report from lymphoscintigraphy and the working definition of ‘nodes to remove’ (e.g. see below for the ‘10% rule’).
If blue dye is used, it can be a useful adjunct for aiding SLN localization and harvesting. Blue dye presents a lower SLN detection rate than radiotracers, but it can be used in addition to radiocolloids. Following injection, the blue dye drains to the SLNs, staining the channels, which can be followed to the first echelon nodes. Direct visualization and dissection of these channels facilitates SLN localization (Fig. 5.9).
FIG. 5.9. Methylene blue dye (1 mL) was injected periareolarly in a 44 year old woman with left breast cancer 10 min prior to the operation. After axillary surgical incision and tissue dissection, a blue stained SLN is clearly seen.
In practice, any LNs that have increased radioactive uptake or vital dye uptake are localized, and more often than not, multiple nodes are detected. The issue of how many SLNs should be biopsied when multiple nodes are found is still being debated. In this regard, while removing too few nodes may miss potential metastases in regional LNs, indiscriminate removal of axillary nodes may cause morbidity similar to that experienced after conventional axillary LDN (in addition to the unnecessarily increased burden for histopathological analysis).
5.4.3.2. SLN non-visualization or failed intraoperative detection
The majority of patients with pre-operative lymphoscintigraphic non-visualization will have at least one SLN detected intraoperatively, either by a gamma probe alone or by a gamma probe combined with blue dye.
Otherwise, a second radiocolloid injection may be useful to depict the previous non-visualized SLN. In approximately 1%–2% of the patients, the SLN will not be detected intraoperatively, and the status of axillary LNs cannot be determined.
Old age, obesity, tumour location other than in the upper outer quadrant and non-visualization of SLNs on pre-operative lymphoscintigraphy may be associated with failed SLN localization [5.83]. The significance of pre-operative scintigraphic SLN non-visualization is not yet known. Some studies have suggested that patients with unsuccessful axillary mapping may have an increased risk of metastatic axillary involvement [5.84]. Current standards of care recommend axillary LN dissection in cases of non-intraoperative SLN identification.
5.4.3.3. Histopathology of SLNs
Detailed histopathological analysis of the SLN is the standard procedure on which to base selection of the postoperative management strategy of breast cancer patients. However, protocols for SLN analysis have not yet been standardized;
therefore, high variability in procedures still exists among different centres.
Different procedures for intraoperative SLN analysis have been developed, including the touch imprint of one or more slices (relatively low sensitivity, but very high specificity), staining of one or several intraoperative frozen sections and even immunohistochemistry for cytokeratins as the most exhaustive method. Immunohistochemistry considerably improves sensitivity by identifying micrometastases and even isolated tumour cells (which are generally missed with conventional haematoxylin and eosin staining alone). Nevertheless, it is still difficult to compare results among different centres, particularly in studies involving detection of micrometastases and isolated tumour cells [5.85].
Molecular biology methods, such as those based on the reverse transcription polymerase chain reaction, are also being used for SLN analysis, although they are generally characterized by relatively poor reproducibility, longer time for intraoperative analysis and their inability to analyse the whole of the LNs. The recently developed one step nucleic acid amplification method is currently being validated in many centres [5.86].
5.5. ADDED VALUE OF INTRAOPERATIVE