Abstract
Background
Sentinel lymph node (SLN) biopsy is a standard procedure in evaluating the status of node negative axilla. Numerous techniques have been described in literature. We hereby describe a new technique of intradermal injection of blue dye called the lymphatic flare technique.
Methods
The study was conducted in two phases over a year from August 2020 to May 2021with an internal audit to validate and standardize the technique in January 2021.
Results
Between August 2020 and December 2020, 32 patients were evaluated for validation of this technique by two senior surgeons, which yielded a SLN identification rate of 93.75% (30 out of 32). After validating, standardizing, and educating the entire surgical team of the technique, another consecutive 27 patients were evaluated. The SLN identification rate increased to 100% (27 out of 27). Overall, SLN positivity for cancer was 16.6% (10 out of 60).
Conclusion
SLN identification by the lymphatic flare technique is feasible, accurate, and reproducible.
Keywords: Sentinel node biopsy, Blue dye, Lymphatic flare
Introduction
Sentinel lymph node (SLN) biopsy is a validated technique to accurately assess the status of clinically negative axilla in early breast cancer which also helps to mitigate the morbidity of an axillary dissection [1]. There are many different techniques of performing SLN biopsy, and all have been validated through multiple trials [2]. The combined blue dye and radioactive colloid–dual tracer technique is recommended in most of the recent guidelines [1, 3, 4]. However, the lack of nuclear medicine facility, concerns regarding radiation exposure, and other logistic issues preclude widespread use of dual tracer, especially in developing countries. Multiple recent studies have reported that blue dye as the single tracer was sufficient for identifying SLNs in breast cancer with SLN identification rates of 91.7% [5, 6].
Here, we describe our novel “lymphatic flare technique” of performing SLN biopsy in carcinoma breast using blue dye which can be easily mastered and practiced widely, especially in resource limited settings such as rural India.
Methods
Consent for sentinel lymph node biopsy was obtained in all patients. All patients with clinical T1 to T2 primary breast tumors (AJCC 8th edition) and clinical-radiologically node negative axilla undergoing upfront surgery were included in the study. Patients who had clinical or radiologically suspicious metastatic nodes in the axilla or if in post-neoadjuvant status, even if node negative axilla, were excluded from the study.
The technique utilizes 1% diluted methylene blue which is injected intradermally in peri areolar location of the breast at 12, 3, 6, and 9 o’clock positions, using a hypodermic insulin syringe (1 ml, 26 G needle). A very small volume of dye (0.2 to 0.3 ml of total dye inclusive of all sites) is injected intradermally so as to raise a small dermal bleb with visualization of blue streaks of dye ramifying from the bleb which we describe as “the lymphatic flare” (Fig. 1). At least three sites of injection should demonstrate a good lymphatic flare to ensure optimal SLN identification. The breast is not massaged; rather a brief, firm, uniform compression is applied over the injected area, and this is followed by gentle massage of the axillary area for less than 30 s. A small incision of length less than 5 cm is placed 1 cm cranial to the lateral breast crease with 1/3 of the incision lying above the anterior axillary line and 2/3 below it in such a way that it corresponds to the lowest axillary hairline. The incision is deepened till the lateral border of the pectoralis major is visualized. The axillary fascia is incised, followed by the clavipectoral fascia along the lateral border of pectoralis minor muscle and thus entering the axillary fossa, where it is possible to visualize blue streaks of lymphatics traversing from chest wall leading to a node which is usually blue stained (Fig. 2). The most common site of identifying the blue node is in zone A of Berg level I of axilla as described by Clough et al. [7], which accounts for 98% of SLN location. This area is anatomically bounded laterally by lateral thoracic vein, medially by chest wall, and cranially by the second intercostobrachial nerve. The blue node (s) or a node at the end of a blue-stained lymphatic channel is harvested along with any other abnormal lymph node in the vicinity. This anatomical knowledge helps in avoiding unnecessary dissection in other zones as described by Clough et al., where the possibility of finding sentinel lymph nodes are rare and thereby decreases the incidence of lymphedema [7].
Fig. 1.
The lymphatic flare
Fig. 2.
Blue sentinel node in zone A
Results
Based on the initial serendipitous observation of association of lymphatic flare and high sentinel node identification rates, a prospective validation study comprising 32 patients between July 2020 and December 2020, wherein the procedure was done by two senior authors (DD and DD). The mean age in this group of patients was 55 years (range from 24 to 83 years). Premenopausal age group comprised 11 patients, and postmenopausal age group had 21 patients. The mean tumor size was 2.98 cm (range from 0.8 to 3.5 cm). The tumor was located most commonly in the upper inner quadrant (UIQ) in 14 patients, followed by upper outer quadrant (UOQ) in 12 patients, lower outer quadrant (LOQ) in 3 patients, lower inner quadrant (LIQ) in 2 patients and one patient with central sector location, respectively. Out of the 32 patients, 28 patients underwent breast conservation surgery (BCS), and the remaining 4 patients underwent mastectomy. The overall SLN identification rate was 93.75% (30 out of 32 patients). The mean number of SLN identified was 1.87 nodes (range from 1 to 5 nodes). The SLN positivity for metastasis was 18.75% (6 out of 32 patients) out of which in one patient the positive node was identified pathologically in the adjacent abnormally enlarged non-blue node (Table 1).
Table 1.
Prospective validated patients between July 2020 and December 2020
| S no | Age | Menstrual status | Tumor size | Location | BCS/mastectomy | SLNB status | No of SLNs | Non-SLN positivity |
|---|---|---|---|---|---|---|---|---|
| 1 | 43 | Pre | 2.5 | UOQ | BCS | Negative | 1 | Negative |
| 2 | 61 | Post | 4 | Central | BCS | Negative | 2 | Negative |
| 3 | 72 | Post | 3.5 | LIQ | BCS | Negative | 1 | Negative |
| 4 | 45 | Pre | 4 | UIQ | BCS | Negative | 1 | Negative |
| 5 | 46 | Pre | 2.5 | UIQ | BCS | Negative | 1 | Negative |
| 6 | 59 | Post | 4.5 | UIQ | BCS | Negative | 3 | Negative |
| 7 | 54 | Post | 5.5 | UOQ | BCS | Negative | 1 | Negative |
| 8 | 24 | Pre | 5 | UIQ | BCS | Negative | 2 | Negative |
| 9 | 73 | Post | 7 | UOQ | BCS | Negative | 2 | Negative |
| 10 | 67 | Post | 2.2 | UOQ | BCS | Negative | 3 | Negative |
| 11 | 81 | Post | 4 | UOQ | BCS | Negative | 2 | Negative |
| 12 | 75 | Post | 2.2 | UOQ | BCS | Not identified | 0 | Negative |
| 13 | 48 | Pre | 1.7 | UOQ | BCS | Positive | 2 | Positive |
| 14 | 29 | Pre | 3.2 | UOQ | BCS | Positive | 1 | Negative |
| 15 | 69 | Post | 2.5 | UIQ | BCS | Positive | 4 | Negative |
| 16 | 49 | Post | 1.5 | LOQ | BCS | Negative | 1 | Negative |
| 17 | 49 | Post | 3.5 | UOQ | BCS | Negative | 3 | Negative |
| 18 | 52 | Post | 2.5 | UIQ | Mastectomy | Negative | 1 | Negative |
| 19 | 73 | Post | 2.6 | UIQ | BCS | Positive | 1 | Negative |
| 20 | 83 | Post | 0.3 | LOQ | BCS | Negative | 1 | Negative |
| 21 | 46 | Pre | 2.2 | UIQ | BCS | Negative | 3 | Negative |
| 22 | 48 | Pre | 5.5 | LIQ | Mastectomy | Negative | 4 | Negative |
| 23 | 51 | Post | 0.8 | UIQ | BCS | Negative | 2 | Negative |
| 24 | 52 | Post | 2.5 | UOQ | BCS | Positive | 1 | Negative |
| 25 | 38 | Pre | 1.5 | UOQ | BCS | Negative | 1 | Negative |
| 26 | 47 | Pre | 1.6 | UIQ | BCS | Negative | 1 | Negative |
| 27 | 65 | Post | 1 | UIQ | Mastectomy | Negative | 2 | Negative |
| 28 | 49 | Post | 2.5 | UOQ | BCS | Not identified | 0 | Negative |
| 29 | 68 | Post | 5.1 | UIQ | Mastectomy | Negative | 5 | Negative |
| 30 | 63 | Post | 3.1 | UIQ | BCS | Negative | 3 | Negative |
| 31 | 33 | Pre | 2.5 | LOQ | BCS | Positive | 2 | Negative |
| 32 | 77 | Post | 2.5 | UIQ | BCS | Negative | 1 | Negative |
Subsequently, the procedure was audited and standardized with respect to the volume of injection, syringe used, technique of injection, minimum number of sites, where lymphatic flare has to be elicited (at least three), time interval between dye injection and axillary incision, need for breast massage, and site of identification of SLN in the axilla. After a brief period of training for the entire surgical team on the standardized procedure [comprising three other surgeons], it was done in another 27 consecutive patients between January 2021 and May 2021. The mean age in this group of patients was 55 years (range from 39 to 86 years). Premenopausal group comprised 10 patients, and postmenopausal group had 17 patients. The mean tumor size was 2.8 cm (range from 1.3 to 4.5 cm). The tumor was located most commonly in UIQ in 10 patients, followed by LOQ in 8 patients, UOQ in 5 patients, LIQ in 3 patients, and one patient with central sector location, respectively. Out of the 27 patients, 26 patients underwent BCS, and the remaining 1 patient underwent mastectomy. The overall SLN identification rate was 100%. The mean number of SLN identified was 2.7 nodes (range from 1 to 5 nodes). The SLN positivity for metastasis was 14.8% (4 out of 27 patients), out of which in 3 patients, the positive node was identified pathologically in the adjacent abnormally enlarged non-blue node (Table 2).
Table 2.
Post-standardization — January 2021 to June 2021
| S no | Age | Mestrual status | Tumor size | Location | BCS/mastectomy | SLNB status | No. of SLNs | Non-SLN positivity |
|---|---|---|---|---|---|---|---|---|
| 1 | 62 | Post | 3.5 | UIQ | BCS | Negative | 1 | Negative |
| 2 | 43 | Pre | 2.5 | UOQ | BCS | Positive | 4 | Positive |
| 3 | 54 | Post | 2.5 | UOQ | BCS | Negative | 1 | Negative |
| 4 | 65 | Post | 3.5 | UOQ | BCS | Negative | 1 | Negative |
| 5 | 45 | Pre | 3 | LIQ | BCS | Negative | 3 | Negative |
| 6 | 65 | Post | 3 | UIQ | BCS | Negative | 4 | Negative |
| 7 | 86 | Post | 1.8 | LOQ | BCS | Negative | 3 | Negative |
| 8 | 50 | Pre | 2.1 | UIQ | Mastectomy | Negative | 1 | Negative |
| 9 | 47 | Pre | 2.2 | UOQ | BCS | Negative | 4 | Negative |
| 10 | 58 | Post | 5.5 | UIQ | BCS | Negative | 3 | Negative |
| 11 | 60 | Post | 1.6 | LOQ | BCS | Negative | 2 | Negative |
| 12 | 69 | Post | 3 | LOQ | BCS | Negative | 3 | Negative |
| 13 | 73 | Post | 2.5 | UIQ | BCS | Negative | 4 | Negative |
| 14 | 47 | Pre | 2.8 | LIQ | BCS | Negative | 1 | Negative |
| 15 | 39 | Pre | 4.2 | LOQ | BCS | Negative | 5 | Negative |
| 16 | 46 | Pre | 3 | LOQ | BCS | Negative | 3 | Negative |
| 17 | 63 | Post | 3 | UIQ | BCS | Negative | 2 | Negative |
| 18 | 69 | Post | 2.5 | UIQ | BCS | Negative | 2 | Negative |
| 19 | 50 | Post | 4.5 | Central | Mastectomy | Positive | 3 | Positive |
| 20 | 67 | Post | 2.5 | UIQ | BCS | Negative | 4 | Negative |
| 21 | 49 | Post | 2.1 | LOQ | BCS | Negative | 1 | Negative |
| 22 | 41 | Pre | 3 | UIQ | BCS | Negative | 1 | Negative |
| 23 | 39 | Pre | 1.5 | LIQ | BCS | Negative | 3 | Negative |
| 24 | 58 | Post | 3.5 | LOQ | BCS | Positive | 3 | Positive |
| 25 | 45 | Pre | 2.5 | UOQ | BCS | Negative | 3 | Negative |
| 26 | 52 | Post | 2.5 | LOQ | BCS | Positive | 5 | Negative |
| 27 | 49 | Post | 3.5 | UIQ | BCS | Negative | 3 | Negative |
In the overall study, including both time periods (total of 59 patients), the SLN identification rate was 96.6% (57 out of 59 patients). In the premenopausal women, the overall SLN identification rate was 95.2% (20 out of 21 patients), while in the post-menopausal group, it was 97.3% (37 out of 38 patients). Elderly (age > 65 years) accounted for 15 patients in the whole study group, and the identification rate was 93.3% (14 out of 15 patients). The overall SLN positivity for metastasis was 16.9% (10 out of 59 patients). There were no adverse events of anaphylactic reactions or injection site skin reactions, and the dye staining of the skin disappears by the third post-operative week.
Discussion
The technique of lymphatic flare was a serendipitous observation, wherein it was noticed that if the lymphatic flare was obtained at the time of intradermal injection, then the axilla showed very clear blue lymphatic streaks which led on to the blue node. It was also noticed that this lymphatic channel delineation was not dependent on breast massage or on the customary practice of waiting for the dye to traverse into the axillary SLN. It was clearly demonstrated in vivo by injecting the dye intradermally with “the lymphatic flare technique” and visualization of the blue dye in less than 30 s in the severed lymphatics of a-priori opened axilla in a few cases.
The explanation of these observations lies in understanding the anatomical architecture of the lymphatics, the chemical properties of dyes and mechanical principles of pressure and flow. Our understanding of the anatomical basis of lymphatic clearance of the breast was based on the seminal works of Sappey in 1870, who described breast lymphatics to be a separate entity from the torso and was constituted by a subareolar plexus and a small number of large lymphatic vessels draining to nodes. Anatomical and functional studies on lymphatics have shown that they start as an avalvular lymph capillary network in the superficial dermis with diameters 20 to 70 μm. They then drain to pre-collectors (70 to 150 μm) in the deep dermis which contains valves. Subsequently, the drainage is to superficial and deep lymphatic collecting vessels (150 to 350 μm) located in the subcutaneous tissue and deep tissue, respectively. These lymphatics are valved and have smooth muscles [8]. Normally the lymphatics are empty, and the lymph is propelled by autonomous rhythmic contractions of lymphangions which contain microliters of lymph. As increased tissue fluid enters the lymphangions, they stretch the walls of the lymphatics and thus incite contractions (Starling’s law) which generates flow [9]. We hypothesize that our technique of lymphatic flare is presumably a visual demonstration of the dye being forced into the dermal reticular lymphatics which is akin to an intra-lymphatic injection. The increased fluid under pressure entering the lymphatics possibly evokes contractions of the vessels due to stretching of the walls and thus generates flow. This autonomic flow is further augmented by the positive pressure on the breast and the negative pressure generated in the axilla by gentle axillary massage which stimulate the lymphatics in the axilla to receive the lymph from the breast which is similar to the principle of manual lymphatic drainage by Földi [10].
Methylene blue is a thiazine dye which has a small molar mass of 320 g/mol which probably helps in gaining easier access to small lymphatics in the dermis and also the reason for its rapid transit. This rapid transit was shown by our in vivo experiment, wherein the dye was noticed at the severed end in a-priori exposed axilla soon after the intradermal injection of the dye by lymphatic flare technique. This explains the avoidance of the customary waiting of 5 to 7 min prior to axillary SLN biopsy which is mandated by the Giuliano technique [11]. Yet another departure from the Giuliano technique is the lack of need for breast massage for 5 min post-injection of the blue dye. Even though manual massage of the breast post-injection of the blue dye has been reported to improve SLN identification, there have been concerns regarding the phenomenon of “benign mechanical transport” [BMT] or tumor spread to SLN caused by massage. A study including 776 early breast cancer patients revealed epithelial cells in the SLN of 9.9% of patients who had pre-SLN biopsy breast massage versus only 3.4% in patients who did not have breast massage [12]. The clinical significance of this finding remains to be investigated further. The lymphatic flare technique obviates the need for breast massage and any concerns of phenomenon like BMT can be definitively laid to rest.
In using a small volume syringe with a narrow gauge needle, the pressure generated in the dermal tissue is far greater than when using a large volume syringe (Bernoulli’s principle), and this may aid in the dye being forced into the dermal lymphatics. According to Bernoulli’s principle, the kinetic velocity of the fluid in a small volume syringe is higher than in a large volume syringe for a similar pressure applied to the plunger.
Notwithstanding these hypotheses, the lymphatic flare technique of SLN consistently produces good identification rates and can be easily learned. This technique by using very low volume of dye [~ 0.2 to 0.4 mL] reduces the incidence of anaphylactic reactions, as shown in a recent meta-analysis [13] which reported that volume of < 2 mL of blue dye and intradermal injection site are both associated with lower rates of anaphylaxis (0.031% and 0.0068%, respectively).
In our study, the first cohort of patients in whom the procedure was done for validation and standardization of the technique, the overall SLN identification rate was 93.75% (30 out of 32 patients). In the subsequent cohort of patients, the SLN identification rate was 100%, which can be explained by the mandatory requirement of at least three sites of intradermal injection showing good lymphatic flare. The overall SLN identification rate was 96.6% (57 out of 59 patients) which is comparable to 96% and 97% SLN identification rate by dual dye technique in ALMANAC and NSABP–B32 trial, respectively. This study also demonstrates superior identification rate of 85.6% using radio isotope or blue dye alone technique in the ALMANAC trial. To compare our study with larger studies is not appropriate, but our study demonstrates the technical feasibility of this technique. A recent meta-analysis by Jiyu Li et al. on SLN mapped by methylene blue dye alone showed a pooled identification rate of 91%, with 89% for peritumoral injection and 94% for peri areolar injection, respectively.
Our study attempts to propose a novel technique of blue dye injection which increases the SLN identification rate. This study is limited by not having a large number of patients, and no head-on comparison with other techniques mentioned in literature. The study was conducted in a single specialized high volume cancer center which is reflected by the high SLN identification rates. However, the relatively short learning curve and easy reproducibility, as shown in the second phase, wherein the rest of the team adapted the technique with superior results, we expect it can be widely adopted by centers across the world.
In the current era of evidence-based medicine, SLN biopsy is part of all standard guidelines for assessing a N0 axilla in carcinoma breast, thereby avoiding unnecessary axillary dissection and its antecedent morbidity. A non-performance of SLN biopsy in indicated cases of early breast cancer and subsequent development of lymphedema has potential for litigation. Our lymphatic flare technique of SLN biopsy can be easily reproduced and validated even in resource limited settings, where the luxury of scintigraphic imaging may not be available in every center across our state or country.
Conclusions
The lymphatic flare technique of SLN biopsy is a serendipitously observed technique and subsequently standardized and internally validated. It consistently produces high identification rates of SLN and low false negativity. This technique is easily reproducible and can be widely applied across centers even where resource or logistics are constrained.
Author Contribution
All contributed equally to the study by performing the procedure, data collection, literature research, and manuscript preparation.
Declarations
Conflict of Interest
The authors declare no competing interests.
Footnotes
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
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