Preoperative Needle Biopsy of Sentinel Lymph Nodes Using Intradermal Microbubbles and Contrast-Enhanced Ultrasound in Patients With Breast Cancer

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1 Women s Imaging Original Research Sever et al. CEUS to Determine SLN Status in Breast Cancer Patients Women s Imaging Original Research Ali R. Sever 1 Philippa Mills 1 Jennifer Weeks 1 Susan E. Jones 2 David Fish 3 Peter A. Jones 2 Willem Mali 4 Sever AR, Mills P, Weeks J, et al. Keywords: axillary lymph node dissection, breast cancer, contrast-enhanced ultrasound, core biopsy, fine-needle aspiration cytology, microbubble contrast agents, preoperative staging, sentinel lymph node biopsy DOI: /AJR Received August 15, 2011; accepted after revision December 16, Department of Radiology, Maidstone Hospital, Hermitage Ln, Maidstone ME16 9QQ, United Kingdom. Address correspondence to A. R. Sever (ali.sever@nhs.net). 2 Department of Surgery, Maidstone Hospital, Maidstone, United Kingdom. 3 Department of Pathology, Maidstone Hospital, Maidstone, United Kingdom. 4 Department of Radiology, University Medical Centre Utrecht, Utrecht, The Netherlands. AJR 2012; 198: X/12/ American Roentgen Ray Society Preoperative Needle Biopsy of Sentinel Lymph Nodes Using Intradermal Microbubbles and Contrast-Enhanced Ultrasound in Patients With Breast Cancer OBJECTIVE. The purpose of this study was to assess whether sentinel lymph nodes (SLNs) that undergo targeted needle biopsy after identification by contrast-enhanced ultrasound (CEUS) using intradermally injected microbubbles results in more node-positive breast cancer patients being diagnosed preoperatively. Furthermore, we sought to determine whether the addition of CEUS to gray-scale sonography of the axilla reduces the number of patients having axillary lymph node (ALN) dissection as a second procedure. SUBJECTS AND METHODS. Intradermal microbubble injection was performed in 136 breast cancer patients who had no abnormal ALNs on routine gray-scale axillary sonography. When an enhancing ALN was visualized, percutaneous sonography-guided fine-needle aspiration cytology or core needle biopsy was performed. Depending on the biopsy results, patients underwent SLN biopsy or ALN dissection. If the putative SLN biopsy was positive or a biopsy tract was seen in the excised SLN, the procedure was defined as successful. RESULTS. SLNs were identified and biopsied in 126 of the 136 cases (93%). Seventeen patients had positive sonography-guided biopsy results (13%) and were treated with immediate ALN dissection. In seven patients, the biopsied node was the only positive node. The remaining 109 patients underwent SLN biopsy. In nine cases (8%), a positive lymph node was identified. Four of these false-negative cases had only micrometastases. CONCLUSION. SLNs can be identified and biopsied using CEUS to increase the accuracy of preoperative axillary staging. If the needle biopsy result is negative, conventional SLN biopsy is indicated. A xillary lymph node (ALN) status provides important staging and prognostic information about patients with invasive breast cancer and may influence treatment decisions [1, 2]. ALN dissection was the gold standard but is associated with significant morbidity [3, 4]. With increased breast cancer awareness, improved imaging systems, and high-quality screening programs, patients now often present with smaller tumors and the incidence of lymph node involvement has decreased. Almost two thirds of breast cancer patients present without axillary metastases [5, 6]. Consequently, sentinel lymph node (SLN) biopsy has replaced ALN dissection as the standard of care for staging the axilla [7, 8]. The SLN is the first lymph node draining the primary tumor, and its histologic status is a reliable indicator of the entire lymph node basin. First pioneered using blue dye in patients with malignant melanoma [9], Giuliano et al. [10] adapted its use in breast cancer and Krag et al. [11] used radiolabeled colloid to identify the SLN. Albertini et al. [12] were the first to use both blue dye and gamma probe guided surgery, leading to an SLN identification rate of more than 95% with false-negative rates between 5% and 10% after subsequent ALN dissection [13]. United States guidelines currently advise completion ALN dissection when the SLN biopsy reveals macro- or micrometastases (> 0.2 mm) [13]. Approximately 50% of patients with positive SLN biopsy findings harbor additional non sentinel ALN metastases. This possibility represents an important drawback of the SLN biopsy procedure because these patients normally require a second operation. More accurate preoperative staging would reduce this need. Clinical examination of the axilla is inaccurate for predicting ALN status [14, 15]. The efficacy of preoperative axillary sonography has been investigated, and the morphologic features of abnormal lymph nodes AJR:198, August

2 Sever et al. have already been described [16]. Particularly when combined with sonography-guided biopsy of abnormal lymph nodes, preoperative axillary sonography has been shown to reduce the need for a second operation in more than 50% of cases [17, 18]. Contrast-enhanced ultrasound (CEUS) is established in clinical practice. IV administration of a contrast agent provides enhanced imaging of tissue microvasculature in many organs. Previously always given IV, microbubbles were initially used mostly for cardiac imaging; their use has been expanded to include hepatic imaging, posttransplant sonography, abdominal trauma scanning, and imaging other areas [19]. Sonographic contrast agents are composed of a dispersion of microbubbles, each of which is smaller than an RBC, that act as reflectors of the ultrasound beam [20, 21]. In 2004, Goldberg et al. [22] showed that microbubbles injected peritumorally in a swine melanoma model were seen to enter lymphatic channels, thus allowing SLNs to be identified accurately using sonography in 90% of cases. After a successful trial of CEUS on a swine model [22] and five patients [23], a validation study was performed. The results of that study showed that this technique can be used to identify the SLN in breast cancer patients [24, 25]. The aim of this study was to investigate the role of preoperative sonography-guided needle biopsy of the enhancing SLN or SLNs after microbubble injection in patients with breast cancer and to determine whether CEUS would be superior to routine grayscale sonography in the preoperative detection of node positivity. Subjects and Methods This prospective study was approved by the local institutional review board and the Medicines and Healthcare Products Regulatory Agency. All patients gave written informed consent. Between November 2009 and December 2010, 136 consecutive consenting patients with breast cancer who were scheduled for primary surgical treatment were recruited into the study. Exclusion criteria were histologically or cytologically proven ALN involvement diagnosed clinically or using standard sonographic imaging and the presence of severe medical comorbidities. All sonographic examinations were performed by a single experienced breast radiologist using an Acuson Sequoia 512 scanner (Siemens Healthcare). This scanner provides conventional gray-scale imaging, pulse-inversion harmonic gray-scale imaging, and contrast-specific sonographic imaging with live dual images of tissue only and contrast agent (Cadence Contrast Pulse Sequencing, Siemens Healthcare). A high-frequency 14-MHz linear-array probe (15L8w, Siemens Healthcare) was used. To reduce microbubble destruction, low-mechanical-index values were applied ( ). All patients with suspected breast carcinoma undergo axillary sonography as part of their initial assessment. Patients with abnormal or suspicious nodes undergo fine-needle aspiration cytology (FNAC) or core biopsy (CB). Rather than FNAC, CB was chosen in some cases when the primary was close to the axilla and in some cases when the node was very enlarged. All patients with normal findings on axillary sonography or negative axillary biopsy results were assessed clinically and patients suitable for routine SLN biopsy were invited to be included in this study. For consenting patients, in order not to delay the surgery, axillary assessment with microbubbles was performed within 1 week of the diagnosis of breast cancer. The microbubble study was also offered to those patients commencing neoadjuvant treatment. The median tumor size was 16 mm (range, mm). Most patients had only a small amount of associated ductal carcinoma in situ. Including the in situ component, the median overall tumor size was 19.5 mm (range, mm). There were six cases of in situ cancer only, 104 cases of invasive ductal cancer, 20 cases of invasive lobular cancer, and six cases of other cancer types. Lymphovascular invasion was seen in 38 cases (28%). The lesions were located in the breast as follows: 76 in the upper outer quadrant, 32 in the upper inner quadrant, 22 in the lower outer quadrant, and six in the lower inner quadrant. The Microbubble Procedure Before microbubble injection, 1 ml of 1% lidocaine was injected into the subcutaneous layer of the areola. The microbubble contrast agent was reconstituted using 2 ml of sterile saline, and the ampoule was shaken vigorously to ensure a homogeneous suspension of bubbles. The ultrasound contrast agent is composed of phospholipid-stabilized microbubbles containing sulfur hexafluoride gas (SonoVue, Bracco Imaging) with a mean diameter of 2.5 μm. Between 0.2 and 0.5 ml of the microbubble suspension was injected into and just under the skin of the areola in the upper outer quadrant using a 1-mL insulin syringe and a 26-gauge needle. Up to three consecutive injections were given intradermally similar to the technique used for radioisotope and blue dye injections in patients undergoing conventional SLN biopsy. A conventional gray-scale ultrasound examination of the axilla was performed before injection of microbubbles, and the positions of the imaged lymph nodes were noted. After injection of the microbubbles, lymphatic channels were visualized immediately on contrast pulse sequencing and were tracked into the axilla. Areas of contrast agent accumulation were then imaged with grayscale or were simultaneously viewed on live dual images to confirm the presence of an architecturally defined lymph node. When no obvious lymphatic filling was seen, the areola area was massaged for seconds and imaging was repeated. If no lymph nodes could be identified after three consecutive injections, the study was abandoned and recorded as a failure. In patients whose nodes were successfully identified, the transit time from injection to arrival in the axillary node was between 15 and 45 seconds. Contrast agent remained in the presumed SLN for up to 3 minutes. Repeating the areola massage often produced a refill of the SLN without the need for a second injection. When an enhancing node was identified in the axilla, a biopsy was performed. The first node to enhance was always biopsied. In most cases there was only one SLN; if two SLNs enhanced simultaneously, both were biopsied. Core biopsy was carried out using a 16-gauge needle and up to three samples were taken and sent for analysis. FNAC was performed with a 21-gauge needle and samples were sent dry on glass slides to cellular pathology where routine H and E staining was carried out followed by cytologic analysis. The entire procedure usually took minutes to perform. Sentinel Lymph Node Biopsy The time between the microbubble study and definitive surgery varied from 5 to 90 days (median, 13 days). Patients with a positive FNAC or CB result underwent ALN dissection. The remaining patients with a negative FNAC or CB result were advised to undergo a standard SLN biopsy using a radioisotope and blue dye. The afternoon before surgery, patients attended the nuclear medicine department for subdermal injection of a radioactive isotope (Nanocoll, GE Healthcare), 40 mbq, in the periareolar upper outer quadrant region. Before surgery and immediately after induction of anesthesia, patients received a 2-mL subdermal injection of blue dye (Bleu Patente V 2.5%, Guerbet) in the periareolar upper outer quadrant region. A standard axillary incision was made, and a gamma probe (Navigator GPS, RMD Instruments) was used to facilitate identification of SLNs. All radioactive lymph nodes that is, all nodes colored blue and, very occasionally, any nodes that looked or felt macroscopically suspicious to the surgeon were excised and sent for standard histologic analysis together with the breast cancer specimen. 466 AJR:198, August 2012

3 CEUS to Determine SLN Status in Breast Cancer Patients TABLE 1: Summary of the 136 Microbubble Procedures Variable Value Total no. of cases 136 Technical failures 10 No lymph nodes identified 4 Inadequate biopsy 4 No biopsy evidence in excised SLNs 2 Technical success, no. (%) of cases 126 (93) True-positive results 17 (13) False-positive results 0 No. of cases treated with SLN biopsy 109 True-negative results 100 False-negative results 9 Sensitivity (%) 65 a Specificity (%) 100 b Positive predictive value (%) 100 c Negative predictive value (%) 92 d Note SLNs = sentinel lymph nodes. a (17 / [17 + 9]). b (100 / [ ]). c (17 / [17 + 0]). d (100 / [ ]). The pathologic assessment of the SLNs was performed according to the Sentinel Lymph Node Biopsy Handbook: New Start guidelines [26]. The pathologist also noted whether there was histologic evidence that the SLN had previously been biopsied by noting the presence of a linear tract of granulation tissue with associated hemorrhage, fat necrosis, or hemosiderin-laden macrophages. Results A total of 136 patients who had a microbubble injection and underwent SLN biopsy or immediate ALN dissection, depending on the results of the procedure, were included in the assessment. There were 135 women and one man with a median age of 62 years (range, years). Of these, 55 patients had screening mammography detected cancers diagnosed by the Maidstone and Tunbridge Wells National Health Service Trust Breast Screening Service and the remaining 81 cases were symptomatic referrals to the breast clinic. There were no adverse events, and a patient survey showed a high level of satisfaction [25]. There were no axillary hematomas found after the axillary needle biopsy. In one patient, microbubbles successfully identified the SLN and the 16-gauge CB was benign, but the surgical SLN biopsy procedure failed to identify any hot or blue nodes. The patient underwent ALN dissection and all nodes were benign. In total, 136 patients underwent SLN biopsy or immediate ALN dissection depending on the results of the preoperative assessment with microbubbles. The microbubble procedure was defined as successful when the following three parameters were met: lymphatic enhancement identified at the site of microbubble injection and followed into the axilla where enhancing lymph nodes were seen; a successful biopsy of these nodes; and pathologic confirmation of the biopsy tract within the excised SLNs. When biopsy of the presumed SLN yielded positive findings, patients proceeded to immediate ALN dissection and these cases were also regarded as successful. The procedure was unsuccessful in 10 patients. In four patients, microbubble injection failed to show any enhancing lymph nodes; therefore, no biopsy was performed. In one of these four cases, the patient had a recent surgical excision and the microbubble contrast agent pooled in the surgical cavity and the lymphatics failed to bypass this area. In two cases the lymphatics were seen but could not be followed into the axilla; in the one remaining case, no lymphatics were identified. In 132 of the 136 cases (97%), a presumed SLN was seen enhancing in the axilla after microbubble injection and a biopsy was performed. CB was performed in 87 patients and FNAC in 83 cases. In 38 cases, both FNAC and CB were performed because of the perceived inadequacy of the FNAC specimen at the time of biopsy. Four biopsies were reported as inadequate because no sufficient cytologic material or lymph node tissue was included within the biopsy specimen. Those cases were also recorded as failures. In 128 cases the biopsy material was adequate. However, in two cases there was no definite histologic evidence of a previous biopsy within the excised SLN, raising the possibility that a lymph node that was not the SLN had been sampled in error. Three of the 10 failed cases (30%) were positive at subsequent SLN biopsy; all were in the group in which microbubble injection failed to show any enhancing lymph nodes. The overall results of the procedure are shown in Table 1. The microbubble procedure was deemed successful in 126 of the 136 cases (93%) when the SLNs were correctly identified and biopsied. In 17 of the 126 cases (13%), histopathologic examination of the needle biopsy sample from the microbubble procedure proved a metastatic deposit in the SLNs and these patients underwent ALN dissection (true-positive biopsy). In seven cases the biopsied node was the only positive node in the axillary specimen (Fig. 1), and in three of those seven cases the only evidence of axillary disease was a micrometastasis ( mm) in the biopsied node. The other 10 patients had more than one involved node in their axillary dissection specimen. In 109 cases there was no evidence of malignancy within the nodal biopsy specimen and these patients underwent SLN biopsy. In nine of the 109 cases (8%), there was malignancy in the excised SLN (false-negative biopsy, Fig. 2). The location of the primary tumor in these nine cases was as follows: seven in the upper outer quadrant, one in the lower outer quadrant, and one in the lower inner quadrant. Eight of the 109 patients (7%) had completion ALN dissection. One patient with micrometastasis declined further axillary surgery. In five of the eight patients (63%) who underwent completion ALN dissection, there were no other involved nodes identified in the axillary specimen. In the remaining three patients, however, the completion ALN dissection specimen contained macrometastatic deposits in the non-slns. In one case, only one of 17 non-slns was positive, containing a 5-mm deposit. The second case had a micrometastasis in the hot and blue SLN containing the biopsy tract and an 8-mm deposit in the other slightly hot, nonblue node, whereas the completion ALN dissection specimen revealed an additional nine of 21 nodes were also involved. In the third case the surgeon removed two SLNs: The hot and blue node was negative and contained a biopsy tract in keeping with the microbubble-enhanced node and the other SLN was nonhot and nonblue but hard and contained an 11-mm metastasis with extracapsular spread. In this patient, the completion ALN dissection specimen showed an additional five of 15 nodes were involved. In most of the cases, the lymphatics were readily followed into the axilla and microbubbles highlighted, by their enhanced reflection, a single axillary node. However, in some cases a second or third node was also noted enhancing subsequently via the efferent lymphatics connecting the first node to the others. In such cases the initial enhancing node was accepted as the true presumed SLN and the remaining nodes were not biopsied. In 16 of the 123 patients (13%) two nodes were identified by simultaneous enhancement and both nodes were biopsied. Three patients in this group were proven to AJR:198, August

4 Sever et al. have axillary metastasis and all were treated with immediate ALN dissection. In 76 of the 136 patients (56%) the enhancing lymph node or nodes were also seen on gray-scale imaging before the microbubble injection. However, in 55 cases (40%) the presumed SLN could be identified as a lymph node by sonography only after the microbubble enhancement. It was otherwise not visible on gray-scale imaging either because of its deep location or because it had a very thin cortex and a prominent fatty hilum and therefore was obscured by the surrounding axillary fat. In five cases (4%), there was a focal collection of contrast agent in the axilla but gray-scale imaging failed to identify a nodal architecture corresponding to the site of enhancement, raising the possibility of contrast pooling rather than true nodal enhancement. Three patients underwent the microbubble procedure before commencing neoadjuvant chemotherapy. Two had a negative SLN FNAC and at surgery had a negative SLN biopsy. The third patient had a positive CB of the presumed SLN after microbubble injection and her final postchemotherapy ALN dissection revealed (residual) isolated tumor cells (< 0.2 mm) in one node. A C Fig year-old woman with grade II invasive ductal carcinoma. A, Contrast-enhanced ultrasound image obtained using contrast-specific mode (Cadence Pulse Sequencing, Siemens Healthcare) reveals nodal enhancement (arrows). B, Conventional gray-scale image shows same node (arrows) as A. C, After sentinel lymph node was identified with microbubbles, core biopsy sample was taken of this node (short arrows) using 16-gauge biopsy needle (long arrows). Discussion SLN biopsy has become the standard procedure for axillary staging in breast cancer patients. However, if metastases are detected in the SLNs, then completion ALN dissection is advised. Preoperative diagnosis of lymph node metastases reduces this need. In most United Kingdom based centers, the routine use of intraoperative assessment of the SLNs with frozen-section or touch imprint cytology has not been adopted for logistical reasons. Many units do not have an onsite histopathology or cytology service. False-negative rates are well recognized for both techniques and the reported patientbased sensitivities are highly variable; a meta-analysis reported a sensitivity of 75% (95% CI, 65 84%) and 63% (95% CI, 57 69%) for frozen-section and touch imprint cytology, respectively [27, 28]. Accurate preoperative axillary staging has many potential benefits. These benefits not only avoid a second operation, but also facilitate better theater scheduling, allow precise patient consenting, and may influence decisions regarding neoadjuvant chemotherapy. In 2004 and more recently in 2011, Goldberg et al. [22, 29] showed that in a swine melanoma model, microbubbles had the potential to identify the sentinel nodes. A subsequent clinical study showed, for the first time, that in breast cancer patients a microbubble contrast agent injected intradermally readily entered breast lymphatic channels and traveled into the draining SLNs and that enhancement was clearly visualized by CEUS [24, 25]. This current study describes how sonography-guided biopsy of the enhancing node has the potential to identify malignancy within otherwise normal-looking SLNs preoperatively and therefore may select patients in whom an SLN biopsy would be futile. Preoperative axillary gray-scale sonography and biopsy of abnormal axillary nodes are becoming routine practice in breast units. A meta-analysis of five studies showed axillary sonography with sonography-guided biopsy to be moderately sensitive (45.4%) and highly specific (99.6%) for the diagnosis of axillary metastases [30]. Others have emphasized the limitations of gray-scale sonography, showing that normal axillary sonography misses involved axillae in 16 29% of cases [31, 32]. These figures are consistent with our audit of data from 2005 to 2008, in which 503 patients with a normal axillary sonography underwent SLN biopsy and B 468 AJR:198, August 2012

5 CEUS to Determine SLN Status in Breast Cancer Patients Fig. 2 Photomicrograph (H and E, 4) of histopathologic slide from sentinel lymph node specimen of 62-year-old woman with grade II invasive ductal carcinoma. Micrometastasis (1.4 mm) was identified within node (arrowheads). Hemorrhage from previous biopsy tract was noted a few millimeters away from metastatic deposit (arrows). in 105 patients (21%) the SLN biopsy result was positive and necessitated a subsequent completion ALN dissection [18]. Since the introduction of the microbubble procedure in our routine practice in 2008, the completion ALN dissection rate has dropped from 21% to 8%. Our experience has clearly shown one of the main limitations of gray-scale sonography is that the SLN can be obscured within the axillary fat. Indeed, we report here that 40% of the SLNs could be identified as a lymph node only after microbubble enhancement. In four of the 17 true-positive (24%) and in five of the nine false-negative (56%) cases, the SLNs could not be seen on gray-scale images alone. These results are also consistent with those of Britton et al. [33]; those investigators found that grayscale sonography guided biopsy missed the SLN in 36% of patients and concluded there was a need for more reliable methods of nodal identification. In experimental studies, Goldberg et al. [22] identified filling defects within enhancing SLNs that were successfully correlated with tumor deposits. In our study, we aimed only to identify and biopsy the SLNs and did not attempt to perform such correlation. Axillary lymph nodes are deeper in location in the human axilla compared with those in their animal models, which makes identification of small filling defects difficult. All patients had an initial assessment of the ipsilateral axilla with clinical examination and sonography. Any abnormal or enlarged nodes were biopsied and those with involved nodes were excluded. In the 26 patients who had positive nodes, none was sonographically enlarged. Recognition of the nodal enhancement pattern may be a valuable addition to this technique and perhaps such filling defects could be better seen in lymph nodes with a thickened cortex. Further studies are required to assess this feature. In the aforementioned experimental studies, both groups used different microbubbles, which may be significant. We have noticed that microbubbles do not last long in the lymph nodes and their persistence is related to the duration of sonography scanning. More robust microbubbles may provide a more homogeneous filling of the lymph nodes and may enhance longer, enabling a more detailed assessment of any filling defects. In patients undergoing neoadjuvant chemotherapy, identification of the SLN using microbubbles may be particularly valuable. Neoadjuvant treatment lowers the rate of SLN identification possibly because of fibrosis within the axilla [34, 35]. In our series, 10 patients underwent neoadjuvant therapy and microbubble assessment. In seven cases the microbubble assessment was performed after completion of neoadjuvant chemotherapy. In three patients the microbubble assessment preceded neoadjuvant chemotherapy. In one of these three patients, a positive node was proven on CB after microbubble assessment and subsequently was found to have only isolated tumor cells in the ALN dissection specimen, indicating a partial response. Although we have no such case in our series of patients with proven metastases on CB after microbubble assessment and later deemed negative after ALN dissection, we acknowledge that this false positivity can occur after a complete pathologic response on axillary nodal disease. In such cases it is possible for the pathologist to identify fibrous scarring in the node where the metastasis had been previously proven. Insertion of a marker into or next to the microbubble-enhancing SLN in these patients may help in identifying the node later if conventional techniques fail. Similarly, patients treated by mastectomy for widespread ductal carcinoma in situ or for risk reduction may benefit from a deployed marker to facilitate subsequent SLN biopsy if any unsuspected invasive component is identified histologically. In four patients microbubble injection failed to show any enhancing lymph nodes, and therefore no biopsy was performed. Three of these four cases had a positive SLN biopsy. It is well established that lymph nodes totally replaced by metastases can be associated with a falsenegative SLN biopsy because these nodes may fail to take up blue dye or radioactive isotope when the lymphatics are obstructed [36]. In such cases microbubble enhancement may also fail. In patients who have no enhancing nodes, cautious gray-scale imaging particularly at the site where abrupt lymphatic visualization stops may help to identify abnormal lymph nodes. Such cases also highlight the importance of having an experienced surgeon who can recognize these involved nodes by the naked eye at operation. A few limitations of this study merit comment. The data are from a single unit and a single radiologist. The procedure has a learning curve not only in identification, but also in the biopsy technique. For this reason we have not included the results of two other radiologists who have recently started performing this technique and validating their results. It is important to differentiate contrast pooling from actual nodal enhancement. In five cases we encountered this problem: An obvious lymph node was not clearly recognizable on the gray-scale images corresponding to the site of enhancement. In four of these cases the biopsy was inadequate, raising the possibility of lymphatic pooling rather than nodal enhancement. This shortcoming was recognized early in the study. Massage frequently overcame the problem; however, if no lymph node was recognizable on gray-scale images after massage, the procedure was regarded as a failure. Another limitation relates to the false-negative biopsy results in nine cases. A similar drawback was recognized by Britton et al. [33] when half of the false-negative cases revealed a biopsy tract within the SLN. It might be possible to overcome this sampling error by removing the entire contrast-identified lymph node for detailed histologic analysis. Breast lesions are increasingly being removed under sonography guidance using vacuum-assisted and other devices. It is possible that similar techniques could be used to remove the entire SLN. This would have the potential to obviate conventional SLN biopsy and replace it with a percutaneous outpatient procedure. In conclusion, SLNs may be identified and biopsied preoperatively using CEUS after injection of microbubbles. Using this technique increases the number of patients proven to have involved ALNs preoperatively, thereby reducing the futile SLN biopsy rate by AJR:198, August

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