单/双酶标记鸡尾酒抗体AMACR/p63/34βE12在前列腺穿刺组织鉴别诊断中的对比观察

前列腺穿刺标本良恶性鉴别诊断是临床病理诊断工作的难点之一。近年来通过应用α-甲酰基辅酶A消旋酶（AMACR）、34βE12、p63抗体分别检测前列腺癌细胞和基底细胞，提高前列腺癌诊断的准确性已得到普遍认同。Jiang等在2005年首先使用混合型AMACR／p63／34βE12抗体和双酶标记法检测前列腺癌，国内也有单酶标记鸡尾酒抗体AMACR／p63／34βE12检测前列腺癌的报道，但单酶标记与双酶标记鸡尾酒抗体两种试剂染色的比较研究尚未见报道。我们采用单酶标记的鸡尾酒抗体和双酶标记的混合型抗体标记前列腺穿刺标本，以评价两种方法在前列腺癌的鉴别诊断中的应用价值。[第一段]     

抗体鸡尾酒AMACR/P63/34βE12在前列腺良恶性病变鉴别诊断中的应用价值

目的:探讨抗体鸡尾酒AMACR/P63/34βE12在前列腺良恶性病变鉴别诊断中的应用价值。方法:收集2001~2005年111例前列腺手术切除标本，其中前列腺腺癌39例，高级别前列腺上皮内瘤（high-grade prostatic intraepithelial neoplasias，HGPIN）29例，非典型性腺瘤样增生（atypical adenomatous hyperplasia, AAH）3例，前列腺结节性增生（benign prostatic hyperplasia, BPH）40例。作抗体鸡尾酒AMACR/P63/34βE12的免疫标记，观察3种抗体在各类病变中的表达情况。结果:39例前列腺腺癌AMACR全部呈阳性，癌巢周围无基底细胞残存（P63/34βE12阴性）。29例高级别前列腺上皮内瘤变，14例（48.3%）腺泡上皮AMACR呈阳性，29例腺泡上皮周围有连续或不连续的基底细胞（P63/34βE12阳性）。3例非典型性腺瘤样增生中2例腺泡上皮AMACR呈弱阳性；3例腺泡上皮周围有较连续的基底细胞（P63/34βE12中度阳性）。40例前列腺结节性增生，腺泡上皮AMACR染色均呈阴性，周围有连续的基底细胞（P63/34βE12强阳性）。结论:鸡尾酒抗体AMACR/P63/34βE12标记前列腺组织，能够同时高特异性和敏感性地检测出前列腺腺癌细胞（或非典型增生的腺泡上皮细胞）和基底细胞，为前列腺腺癌与高级别上皮內瘤变、非典型性腺瘤样增生、前列腺结节性增生的鉴别诊断提供有力的证据。     

单/双酶标记鸡尾酒抗体AMACR/p63/34βE12在前列腺穿刺组织鉴别诊断中的对比观察

前列腺穿刺标本良恶性鉴别诊断是临床病理诊断工作的难点之一.近年来通过应用α-甲酰基辅酶A消旋酶(AMACR)、34βE12、p63抗体分别检测前列腺癌细胞和基底细胞,提高前列腺癌诊断的准确性已得到普遍认同[1～5].Jiang等[6]在2005年首先使用混合型AMACR/p63/34βE12抗体和双酶标记法检测前列腺癌,国内也有单酶标记鸡尾酒抗体AMACR/p63/34βE12检测前列腺癌的报道[7,8],但单酶标记与双酶标记鸡尾酒抗体两种试剂染色的比较研究尚未见报道.我们采用单酶标记的鸡尾酒抗体和双酶标记的混合型抗体标记前列腺穿刺标本,以评价两种方法在前列腺癌的鉴别诊断中的应用价值.     

混合型p504s/34βE12/p63抗体在前列腺腺癌诊断中的应用价值

目的评价混合型p504s/34βE12/p63抗体和双重染色技术在诊断前列腺腺癌中的价值。方法选择47例前列腺病例,包括36例前列腺癌、4例不典型腺瘤样增生、4例良性增生和3例间质肉芽肿性炎。采用混合型p504s/34βE12/p63抗体和双酶标记法检测这3种抗原在各类病变中的表达状况。p504s阳性表现为胞质内蓝黑色颗粒状物,而基底细胞的核(p63)和胞质(34βE12)呈红色着色。结果在36例前列腺癌中,32例(88·89%)表达p504s,其中5例为小灶性癌,所有癌细胞巢周围无肌上皮细胞(34βE12和p63阴性)。在4例不典型腺瘤样增生中,1例p504s阳性,但腺上皮细胞周围也有肌上皮细胞(34βE12和p63阳性)。其余7例良性病变中p504s阴性。无非癌性病变同时出现p504s阳性而34βE12和p63阴性的情况。单标抗体p504s和混合型p504s在前列腺病变中的表达一致率为91·49%。结论混合型p504s/34βE12/p63抗体双标在前列腺癌的诊断中具有很高的敏感性和特异性,与单标抗体比较,更容易观察、分辨,有助于提高小灶性前列腺癌的检出率。该法适用于常规病理诊断中,尤其是穿刺活检的小标本,而且减少了免疫组化染色片的数量。     

假增生型前列腺癌的病理学特征

目的探讨假增生型前列腺癌（PHPA）的临床病理特征及其发生率和生物学行为。方法复查上海交通大学附属第六人民医院2005年1月1日-2006年12月31日860例直肠B超引导下前列腺穿刺活检和46例前列腺癌根治手术切片,对疑有PHPA组织作34βE12（或CK5／6）、p63和AMACR单项免疫组织化学标记（EnVision法）和34βE12／p63／AMACR鸡尾酒抗体双重免疫组织化学标记,将在1个组织块中PHPA占该组织块中癌总量的面积百分比〉60％的病例归入本组,并作病理学分析。结果PHPA在穿刺活检和前列腺癌根治标本中的发生率分别为7．0％和15．2％。66．7％的PHPA与普通型前列腺癌直接移行,76．7％在其他组织块中有普通型癌。PHPA占穿刺活检中癌总量的比例为5％～100％,占根治标本中癌总量的比例为1％～30％。PHPA以大中腺泡增生为主,癌细胞分化较好,排列有极性,腔内常有残存淀粉样小体,低倍镜下类似良性前列腺增生。但腺泡排列紧密,腔内有嗜酸性结晶体和颗粒状无定形物质,核增大,有大核仁,免疫标记AMACR阳性,基底细胞标记阴性,在20项提示恶性的形态学指标中10项出现几率966．7％。PHPA虽然分化较好,但66．7％的PHPA有间质浸润,6．7％有神经浸润,3．3％有腺外浸润,3．3％发生骨转移,肿瘤分布部位周围带多于移行带。结论PHPA的实际发生率不低,绝大多数与普通型癌并存,由于形态学类似良性,肿瘤细胞量又不占多数,因此在诊断中容易被忽视,PHPA高分化前列腺癌不同,应属于Gleason3级的中分化腺癌。     

前列腺导管腺癌和Gleason4筛状与非筛状腺泡性腺癌的免疫组化抗体鸡尾酒染色

AMACR的过度表达与基底细胞标记（如p63和高分子量角蛋白HMWCK）的缺乏是经典型前列腺腺泡性腺癌的特点。我们利用p63／HMWCK／AMACR鸡尾酒免疫组织化学染色方法，以研究其在前列腺导管腺癌和Gleason评分为4分的筛状腺泡性腺癌中的表达以及在诊断中的应用，并与Gleason评分为4分的非筛状腺泡性腺癌进行比较。这一研究包括了62例前列腺切除标本，其隐含前列腺导管腺癌（n=51）、Gleason4的筛状腺泡性腺癌（n=27）和Gleasorl4的非筛状腺泡性腺癌（n=48），具有代表性的档案蜡块1～4个，均经福尔马林固定，石蜡包埋。     

前列腺导管腺癌42例临床病理分析

目的探讨前列腺导管腺癌的临床病理和免疫组化特征。方法回顾性分析42例前列腺穿刺活检、经尿道前列腺切除和前列腺癌根治手术标本中的前列腺导管腺癌,所有病例均作34βE12、CK5/6、p63、AMACR、PSA和PAP免疫标记,并对照HE切片诊断。结果导管腺癌以周围型多见(39例,92.6%),有30例(21.4%)合并普通经典型腺癌。镜下以大腺泡为主,呈乳头状,筛孔状或管状结构,瘤细胞高柱状,核异型性明显。免疫组化表型类似经典型腺癌,但有23.8%的病例肿瘤性腺管周围有34βE12、CK5/6、p63标记阳性的基底细胞存在。结论导管腺癌与经典型腺癌相比,临床病理和免疫组化表现均有差异,病理诊断应注意与高级别上皮内瘤和转移性腺癌鉴别。     

前列腺不典型小腺泡增生的病理形态及临床意义

目的 探讨前列腺不典型小腺泡增生的形态学特点和临床意义。方法 收集解放军总医院病理科2004-2006年前列腺穿刺活检诊断为不典型小腺泡增生病例11例,复习HE和免疫组织化学切片,并对有不典型小腺泡增生病变的蜡块重新进行34βE12、p63和P504S免疫组织化学（SP法）染色,观察不典型小腺泡增生的组织学特点和免疫组织化学表达特点。结果 11例不典型小腺泡增生均表现为排列紧凑的小腺体,其中6例小腺泡数量在3个以下,圆形或轻度不规则形,核呈单层排列,有的细胞核之间间隔较大。细胞核普遍增大,圆形或不规则形,部分可见明显的核仁。胞质呈嗜双色性或空亮,腔缘相对平整,部分腔内可见蓝色黏液。免疫组织化学显示34βE12、p63阴性,P504S阳性或弱阳性。4例腺泡数量超过3个,圆形或轻微不规则形,细胞核轻度增大,核仁不清楚或有小核仁。34βE12及p63阴性或点状阳性,P504S弱阳性或阴性。11例患者二次穿刺活检诊断为癌的有4例,多为第一次活检中腺泡数量较少但有明确细胞异型性的病例。结论不典型小腺泡增生是一种与前列腺癌密切相关的病变,其腺体数量或细胞形态或组织结构改变不足以诊断为癌的一类病变。不典型小腺泡增生病例二次活检发现癌的几率明显高于一般的增生。     

α-甲酰基辅酶A消旋酶在前列腺腺癌诊断和鉴别诊断中的作用

目的观察α-甲酰基辅酶A消旋酶(α—methylacyl—CoA racemase,P504S)在前列腺腺癌的诊断和鉴别诊断中的价值。方法对前列腺腺癌及其需要鉴别的病变：前列腺上皮内瘤变、不典型腺瘤样增生、不典型小腺泡增生以及正常前列腺组织(包括萎缩的腺体)和良性增生进行光镜观察,用EnVision二步法免疫组织化学方法检测P504S、细胞角蛋白(CK)34βE12、p63在各类病变组织中的表达情况。根据P504S阳性表达部位区分其表达形式为：胞质型、腔缘型、顶端胞质型及胞膜型。结果78例前列腺腺癌中,91％(71／78)阳性表达P504S,多表现为癌细胞弥漫性胞质内阳性着色较深的颗粒状物,少数为腺腔内缘或顶端胞质内及膜表达,9％(7／78)阴性表达P504S者均为亮细胞型;前列腺上皮内瘤变(9例),不典型腺瘤样增生(6例)以及不典型小腺泡增生(2例)中均见P504S不同程度的表达;96％(65／68)的正常及良性增生前列腺组织未见P504S阳性表达;增生的基底细胞也未见P504S阳性表达。结论P504S的免疫组织化学染色对判断前列腺腺癌具有重要参考价值,若与CK34βE12或p63联合应用则更有帮助。     

α-甲酰基辅酶A消旋酶诊断前列腺癌的敏感性和特异性研究

随着α-甲酰基辅酶A消旋酶（AMACR,P504S）在前列腺癌鉴别诊断中的广泛应用,其问题也逐渐显现。我们采用AMACR／34βE12／p63双重免疫组织化学染色技术对AMACR的非特异性表达以及治疗后和各种特殊类型前列腺癌的表达作了较系统的研究。[第一段]     

Using an AMACR (P504S)/34betaE12/p63 cocktail for the detection of small focal prostate carcinoma in needle biopsy specimens

We assessed the usefulness of immunohistochemical analysis with a 3-antibody cocktail (alpha-methylacyl coenzyme A racemase [AMACR, or P504S], 34betaE12, p63) and a double-chromogen reaction for detection of limited prostate cancer in 138 needle biopsy specimens, including 82 with small foci of prostatic adenocarcinoma and 56 benign prostates. When carcinoma was present, red cytoplasmic granular staining (AMACR) in the malignant glands and cells and dark brown nuclear (p63) and cytoplasmic (34betaE12) staining in basal cells of adjacent nonmalignant glands were found. Of 82 cases of small foci of prostatic adenocarcinoma, 78 (95%) expressed AMACR; all malignant glands were negative for basal cell staining. All benign glands adjacent to malignant glands were recognized easily by basal cell marker positivity and little or no AMACR expression. No benign glands were simultaneously positive for AMACR and negative for basal cell markers (specificity, 100%). There were no differences in intensity and numbers of positive glands with double-chromogen staining compared with using 1-color staining. Our results indicate that immunohistochemistry with a 3-antibody cocktail and double chromogen is a simple and easy assay that can be used as a routine test, which overcomes the problems of studying small lesions in prostate needle biopsies with multiple immunohistochemical stains.     

Usefulness of basal cell cocktail (34betaE12 + p63) in the diagnosis of atypical prostate glandular proliferations

We evaluated the diagnostic usefulness of the 34betaE12-p63 cocktail, compared with 34betaE12 and p63 used alone, in 34 prostate needle biopsy (NBXs) and 3 transurethral resection specimens containing atypical glandular proliferations and in 18 NBXs containing unequivocal prostate carcinoma (PCa). Staining intensity; percentage of basal cells staining in benign, atypical, and malignant glands; number of benign glands lacking basal cell staining; and staining variance were analyzed. All NBXs with unequivocal PCa were negative with all 3 markers. Diagnoses were as follows for the atypical cases after staining for the 3 markers: PCa, 9; postatrophic hyperplasia, 12; high-grade prostatic intraepithelial neoplasia (HGPIN), 5; atypical adenomatous hyperplasia, 6; benign atypical proliferations, 4; and HGPIN with adjacent small atypical acinar proliferation suggestive of PCa, 1. The cocktail demonstrated consistently strong staining intensity and improved basal cell staining in morphologically benign and benign atypical glands compared with p63 and 34betaE12 alone; it had the smallest staining variance compared with 34betaE12 (F < 0.0001) and p63 (F = 0.31), although its advantage for resolving individual atypical cases was limited compared with 34betaE12 and p63 alone. Of 37 atypical cases, 1 (3%) additionally was resolved as benign using the cocktail and p63. Because the diagnosis of PCa is supported by lack of basal cell staining, the immunohistochemical analysis with highest possible sensitivity and lowest possible variability is critical to ensure that a negative reaction is true. The cocktail provides a simple, cost-effective improvement in basal cell immunohistochemical analysis of difficult prostate lesions.     

Alpha-methylacyl-CoA racemase (P504S)/34betaE12/p63 triple cocktail stain in prostatic adenocarcinoma after hormonal therapy

Alpha-methylacyl-CoA racemase (AMACR) has recently been shown to be a highly sensitive marker for the diagnosis of prostate cancer. However, there is limited information concerning its utility as a marker for prostate carcinoma after hormonal therapy. Our current investigation was conducted to evaluate the expression of AMACR in patients with prostate carcinoma after hormonal therapy and assess its diagnostic utility in combination with p63 and high molecular weight cytokeratin (34betaE12) staining. Prostate tissues from 49 patients who had been treated with hormonal therapy were immunohistochemically analyzed for AMACR, 34betaE12, and p63 expression by a triple antibody cocktail stain. The staining intensities and the percentages of positively staining tumor cells were recorded. The correlations between AMACR expression and metastatic status, associated hormonal therapy regimens, and the extent of hormone therapy effect were analyzed. All malignant acini were completely negative for both basal cell markers (34betaE12 and p63). Tumor cells failed to demonstrate expression of AMACR in 14 (29%) of 49 cases. In the remaining 35 cases (71%), positive immunostaining for AMACR was noted, but with variable intensities and percentages of cells stained. Positive staining for AMACR in benign glands was not seen in any case. In all cases, basal cells were strongly stained by p63 in benign acini with a mean positive percentage of 96%. Similarly, basal cells in benign acini displayed moderate staining intensities for 34betaE12 in 3 (7%) of 41 cases and strong immunostaining for this marker in the remaining 38 cases (93%); the mean percentage of positive cells was 92%. alpha-methylacyl-CoA racemase expression may be substantially diminished or entirely lost in prostate carcinoma after hormonal therapy. This variation in AMACR expression does not correlate with the metastatic status, the modality of hormonal therapy, or the extent of therapy-related effect. It is important that pathologists be aware that some hormonally treated prostate carcinomas do not express AMACR, and that immunostaining in such cases must be interpreted with caution. A triple cocktail stain using AMACR, 34betaE12, and p63 can be helpful in evaluating prostate specimens for the presence of residual or recurrent carcinoma after hormonal therapy for cancer.     

Is triple immunostaining with 34betaE12, p63, and racemase in prostate cancer advantageous? A tissue microarray study

This study aimed to determine the usefulness of a combination of 3 immunohistochemical markers, 34/betaE12, p63 and alpha-methylacyl coenzyme A racemase (AMACR), for the diagnosis of prostate cancer using tissue microarrays (TMAs) constructed from 91 archival radical prostatectomy specimens derived from the Pathology Department files of Singapore General Hospital, Singapore. Triple immunostaining using a cocktail of these 3 antibodies was performed on TMA sections using the streptavidin-biotin method. When compared with immunohistochemical staining using the individual antibodies, we found that the triple cocktail allowed improved evaluation of basal cells in benign glands, and AMACR allowed simultaneous corroboration of malignant prostatic glands in the same section. We achieved a specificity of 100% with the triple cocktail, and sensitivity was acceptable, at 93.8%. In comparison, specificity and sensitivity of the individual antibodies were 95.5% and 97.3%, 93.3% and 93.8%, 97.0% and 95.6% for p63, 34betaE12, and AMACR, respectively. The triple cocktail offers a cost-effective way of evaluating abnormal prostatic glandular foci, in addition to maximizing the use of small tissue samples from prostatic needle biopsies.     

Alpha-methylacyl coenzyme A racemase immunoreactivity in partial atrophy of the prostate

It was reported that 15 of 19 consultation cases of prostatic partial atrophy were a-methylacyl coenzyme A racemase (AMACR)-positive. We investigated partial atrophy cases from a single institution using a standard AMACR immunostaining method. Immunohistochemical analysis was performed using an antibody cocktail containing p63, high-molecular-weight keratin (34bE12), and AMACR antibodies on 122 foci of partial atrophy. AMACR staining was analyzed in partial atrophy (n = 122) and compared with adjacent benign glands (n = 122) and prostatic carcinomas (n = 28). Of 122 foci of partial atrophy, 38 (31.1%) showed AMACR immunoreactivity. Typically, AMACR staining was weak or moderate. In addition, 82 (67.2%) showed patchy to negative distribution of basal cells. Partial atrophy may show AMACR immunoreactivity when using a 34bE12, p63, and AMACR antibody cocktail staining method. Compounding this problem, focal lack of basal cells may be seen. However, the AMACR staining pattern of partial atrophy is usually comparable to that of adjacent benign glands and substantially different from adenocarcinoma.     

Stratified epithelium in prostatic adenocarcinoma: a mimic of high-grade prostatic intraepithelial neoplasia.

Typically glands of prostatic adenocarcinoma have a single cell lining, although stratification can be seen in invasive carcinomas with a cribriform architecture, including ductal carcinoma. The presence and diagnostic significance of stratified cells within non-cribriform carcinomatous prostatic glands has not been well addressed. The histomorphological features and immunohistochemical profile of cases of non-cribriform prostatic adenocarcinoma with stratified malignant glandular epithelium were analyzed. These cases were identified from needle biopsy cases from the consultation files of one of the authors and from a review of 150 consecutive in-house needle biopsy cases of prostatic adenocarcinoma. Immunohistochemistry was performed utilizing antibodies reactive against high molecular weight cytokeratin (34betaE12), p63 and alpha-methylacyl-coenzyme-A racemase (AMACR). A total of 8 cases were identified, including 2 from the 150 consecutive in-house cases (1.3%). In 4 cases, the focus with glands having stratified epithelium was the sole carcinomatous component in the biopsy, while such a component represented 5-30% of the invasive carcinoma seen elsewhere in the remaining cases. The main attribute in all these foci was the presence of glandular profiles lined by several layers of epithelial cells with cytological and architectural features resembling flat or tufted high-grade prostatic intraepithelial neoplasia, but lacking basal cells as confirmed by negative 34betaE12 and/or p63 immunostains in all cases. The AMACR staining profile of the stratified foci was variable, with 4 foci showing positivity, and 3 foci being negative, including two cases that displayed AMACR positivity in adjacent non-stratified prostatic adenocarcinoma. Prostatic adenocarcinoma with stratified malignant glandular epithelium can be identified in prostate needle biopsy samples harboring non-cribriform prostatic adenocarcinoma and resembles glands with high-grade prostatic intraepithelial neoplasia. These 'PIN-like' carcinomas can present in pure form. Recognition of this pattern of prostatic adenocarcinoma is necessary to correctly diagnose such cases as invasive carcinoma.     

Detection of prostate cancer by alpha-methylacyl CoA racemase (P504S) in needle biopsy specimens previously reported as negative for malignancy

AIMS : To investigate the possibility of detecting small focal prostatic cancer by alpha-methylacyl CoA racemase (AMACR)/P504S immunohistochemistry on needle biopsy specimens that were previously interpreted as negative for carcinoma on routine haematoxylin and eosin (H&E)-stained sections. METHODS: Prostate needle biopsy specimens (n = 793) previously interpreted as benign prostatic tissue by conventional morphology from 239 patients with prostatic cancer diagnosed in other biopsy cores taken at the same biopsy session were stained with the P504S monoclonal antibody. If a biopsy specimen stained positively, two pathologists independently reviewed the original corresponding H&E-stained sections to establish the malignant diagnosis. RESULTS: Eighty-four of the 793 biopsy specimens showed AMACR immunoreactivity; nine of these (9/793, 1.1%) contained previously unrecognized small focal prostatic carcinoma (Gleason 6, N = 8; Gleason 8, N = 1). Six of nine (67%) carcinomas showed foamy/pseudohyperplastic (N = 3) or atrophic (N = 3) features. Additionally, five biopsy specimens (5/793, 0.6%) with positive AMACR staining that did not meet the criteria for prostatic cancer on the original H&E slides were considered to be atypia. CONCLUSIONS: In this study, we found a 1.1% false-negative rate for carcinoma on routine H&E-stained sections. AMACR immunohistochemical staining has shown the ability to improve detection of small focal prostatic carcinoma that could be missed by conventional histological examination.     

Prospective evaluation of AMACR (P504S) and basal cell markers in the assessment of routine prostate needle biopsy specimens

Distinguishing benign prostate glands from malignant ones, based purely on morphology, on prostatic core needle biopsy specimens (PNBs) may prove difficult, particularly if the suspicious focus is small. In recent years, several immunohistochemical markers, including the basal cell cocktail (BCC), 34betaE12 and p63, and the prostate cancer (PCa) biomarker alpha-methylacyl-CoA-racemase (AMACR), have been used as adjuvants to morphology, in these diagnostically challenging cases. We prospectively address the diagnostic utility of using the BCC, in combination with the commercially available AMACR monoclonal antibody, P504S, on PNBs that required immunohistochemistry (IHC) studies to make a diagnosis. The goals of this prospective study were to assess the day-to-day practice in an academic setting, to determine how often these IHC tests were used on routine PNBs, and to establish how often a combination of the BCC and P504S were helpful in diagnosing prostate cancer. A total of 772 prospectively collected PNB cases were examined over a 7-month period. IHC staining was performed in 171 cases (22%); 123 cases were stained with the BCC in addition to the commercially available monoclonal AMACR antibody. In 86 of these 123 cases (70%), both stains contributed to the final diagnosis: PCa in 44 cases, benign in 33 cases and high-grade prostatic intraepithelial neoplasia in 9 cases. Of the remaining 37 cases (30%), 18 were called benign or PCa, based solely on appropriate staining with the BCC, with AMACR being noncontributory because the focus of interest had been cut through (12 cases), there was negative staining with AMACR (in 4 PCa cases), or there was positive staining with AMACR (in 2 benign cases showing atrophy). Nineteen of 37 cases were diagnosed as atypical small acinar proliferation. In these 19 cases either the focus had been cut through on one or both of the stains (11 cases), both AMACR and BCC failed to work (2 cases), AMACR was positive in the presence of patchy BCC staining (1 cases), AMACR was negative in the absence of BCC staining (3 cases), or despite appropriate staining the focus consisted of 1 gland and was considered too small to call carcinoma (2 cases). Additional IHC stains were performed in 171 of 772 cases; of these, 123 had sufficient material to perform both the BCC and P504S. The BCC when used in combination with AMACR rendered a diagnosis in almost 70% of cases. Using these stains in combination may be a better approach in diagnostically difficult cases as it increases the likelihood that a definitive diagnosis can be rendered while decreasing the likelihood of an equivocal diagnosis. However, a limitation of this approach is the loss of tissue in these small lesions, suggesting that combining AMACR and the BCC on a single slide would be superior to using either marker separately.