【正文】 increased signal following deglycosylation, 呈增加的信號(hào)以下deglycosylation ， which is consistent with the notion that the antigenic deter 這是與抗原的概念一致阻止 minant is in close proximity to a glycosylation site in SHBG minant是在SHBG的糖基化位點(diǎn)靠近 and that carbohydrate removal allows antibody 7H9 im 和碳水化合物的去除允許抗體7H9 IM proved access to its binding site. 證明其結(jié)合位點(diǎn)的訪問 。 The efficiency of degly 效率degly cosylation was confirmed by parallel treatment of bovine cosylation證實(shí)了并行處理的牛 fetuin, which showed a similar migration shift on SDS 胎球蛋白，這顯示了類似的遷移轉(zhuǎn)變的SDS PAGE (data not shown). 頁(yè)（數(shù)據(jù)未顯示）。 The relative antigenic determinant mapping experiments 相對(duì)的抗原決定簇映射實(shí)驗(yàn) show that 11F11 monoclonal antibody (isotype IgG2a) does 表明11F11的單克隆抗體（IgG2a亞型 ） not share antigenic determinants with any of the three IgG1 不與任何三個(gè)IgG1的共享抗原決定 monoclonal antibodies (2G11, 7H9, or 16D5) (Fig. 2). 單克隆抗體（2G11，7H9，或16D5）（圖2）。 Comparisons in the upper panel of Fig. 在上游面板圖進(jìn)行比較 。 2, with respect to 2，對(duì)于 antibody 2G11, show that the doseresponse curve gener 抗體2G11，表明產(chǎn)生的劑量反應(yīng)曲線 ated when Dako antibodybound SHBG was probed with ated時(shí)Dako公司抗體綁定SHBG探測(cè)與 antibody 2G11 followed by 11F11 and finally antimouse 11F11和抗體2G11終于antimouse IgG2aperoxidase is identical to that with 11F11 followed IgG2a過(guò)氧化物酶是相同的，11F11其次 Table 4 表4 Linearity of plasma samples from 3 patients (samples 1, 2, and 3) assayed for SHBG at various plasma dilutions using 11F11/2G11 and 11F11/16D5 從3例（樣品1，2，3）血漿樣品線性測(cè)定SHBG在不同的血漿稀釋，使用11F11/2G11和 11F11/16D5 antibody binations 抗體組合 Sample 樣品 Antibody bination 抗體結(jié)合 11F11/2G11 11F11/2G11 Antibody bination 抗體結(jié)合 11F11/16D5 11F11/16D5 1 1 2 2 3 3 1 1 2 2 3 3 Dilution 稀釋 1:1000 1:1000 5 5 2 2 64 64 5 5 192 192 20 20 10 10 2 2 62 62 5 5 242 242 21 21 1:2000 1:2000 4 4 2 2 28 28 3 3 72 72 6 6 7 7 2 2 27 27 4 4 135 135 10 10 1:4000 1:4000 a 一 14 14 2 2 33 33 4 4 a 一 13 13 2 2 74 74 6 6 1:8000 1:8000 a 一 7 7 2 2 16 16 2 2 a 一 5 5 1 1 33 33 4 4 Values (nmol/l) are means of duplicates SD. 值（nmol / L的）是重復(fù)的SD。 a 一 Not determined. 未確定。 263 263 JG Lewis et al. JG劉易斯等人。 / Steroids 64 (1999) 259–265 /類固醇64（1999）259265 Page 6 第6頁(yè) by antimouse IgG2aperoxidase, suggesting no interference antimouse IgG2a過(guò)氧化物酶，提示無(wú)干擾 by antibody 2G11. 抗體2G11。 This is confirmed by the identity of 這是確認(rèn)身份 curves using antibodies 2G11 followed by 11F11 and anti 曲線11F11和抗抗體2G11 mouse IgG1peroxidase and antibody 2G11 followed by 鼠IgG1過(guò)氧化物酶和抗體其次2G11 antimouse IgG1peroxidase. antimouse IgG1的過(guò)氧化物酶。 Also noteworthy was the lack 另外值得一提的是缺乏 of response using antibody 11F11 followed by antimouse 響應(yīng)使用antimouse其次的抗體 11F11 IgG1peroxidase as a control on isotype specificity. 作為控制亞型特異性IgG1的過(guò)氧化物酶。 Simi 西米， larly antibodies, 2G1, 7H9, and 16D5, when followed by larly抗體，2G1，7H9，16D5，其次 antimouse IgG2aperoxidase showed no response (data not antimouse IgG2a過(guò)氧化物酶顯示沒有響應(yīng)（數(shù)據(jù)未 shown). 所示）。 Comparison of the Dako polyclonal SHBG ELISA to the Dako公司多克隆SHBG ELISA相比， monoclonal antibodybased SHBG ELISA for the 11F11/ 單克隆抗體為基礎(chǔ)的SHBG ELISA 11F11 / 16D5 bination and the 11F11/2G11 bination are 16D5組合和11F11/2G11結(jié)合 shown in Fig. 如圖。 3a and b, respectively. 分別為3A和B ， These had been de 這些已被取消 termined by using various binations of Protein Apuri 通過(guò)使用各種組合的一個(gè)，純化蛋白 termined fied SHBG monoclonal antibodies and indicated that opti fied性激素結(jié)合球蛋白單克隆抗體，并表示，OPTI mal results were obtained with 11F11 as the coating 正常結(jié)果獲得11F11涂層 antibody and either 16D5 or 2G11 as the SHBG secondsite 抗體和16D5或SHBG第二現(xiàn)場(chǎng)的 2G11 antibodie. antibodie。 A later, second series of experiments using dif 后來(lái)，第二個(gè)系列實(shí)驗(yàn)使用不同 ferent patient samples showed a parison of the mono ferent病人樣本顯示一個(gè)比較單 clonal antibodybased SHBG ELISAs with the ligand bind 單克隆抗體為基礎(chǔ)的SHBG酶聯(lián)免疫吸附與配體綁定 ing method, Fig. ING方法，圖。 3c and d. 3C和D. SHBG determinations on deglycosylated plasma and deglycosylated血漿和SHBG測(cè)定 controls are shown in Table 1. 控制見表1。 SHBG determinations on SHBG測(cè)定上 partially purified and deglycosylated SHBG pared to 部分純化和deglycosylated SHBG相比 controls are shown in Table 2. 控制表2所示。 Together they provide sub 它們共同提供分 stantial additional information regarding the nature of the stantial有關(guān)的其它信息的性質(zhì) binding sites on SHBG by the various antibodies. SHBG的各種抗體的結(jié)合位點(diǎn) 。 They 他們 confirm that the antigenic determinant of 11F11 is not 確認(rèn)的11F11的抗原決定不 mon with the other monoclonal antibodies and, like 共同與其他單克隆抗體，就像 2G11, is relatively unaffected by carbohydrate removal. 2G11，相對(duì)碳水化合物去除的影響 。 With respect to antibody 7H9 the results showing signifi 關(guān)于抗體7H9的結(jié)果呈現(xiàn)出顯著 cantly elevated SHBG levels following carbohydrate re cantly升高SHBG水平以下碳水化合物重新 moval are consistent with the Western blotting data and 去除率都與免疫印跡數(shù)據(jù)一致 suggest enhanced binding following carbohydrate removal. 建議增強(qiáng)約束力以下的碳水化合物去除 。 This is also the case with antibody 16D5, where even higher 這也是與抗體16D5的情況下，甚至更高 apparent levels of SHBG were measured following carbo 明顯的SHBG水平測(cè)定碳 hydrate removal. 水合物去除。 It, therefore, seems likely that the SHBG 因此，它似乎在SHBG recognition sites by 7H9 and 16D5 are in close proximity to 7H9和16D5識(shí)別位點(diǎn)是在接近 either N or Olinked oligosaccharides and that their re N或O型掛鉤低聚糖和他們重新 moval allows improved antibody access. 去除率允許提高抗體訪問。 When another se 當(dāng)另一SE rum sample was subjected to Olinked oligosaccharide re 糖酒會(huì)樣品是O型掛鉤低聚糖重新 moval, which includes a desialylation step, and assayed 去除率，其中包括desialylation步驟，和檢測(cè) using immobilised 11F11 and either 7H9 or 16D5 as the 使用固定11F11和7H9或作為16D5 detecting antibody, there were no differences between the 檢測(cè)抗體，之間有沒有差異 enzyme treated and control samples for each antibody, sug 酶處理和對(duì)照樣品，每個(gè)抗體， SUG gesting that the SHBG recognition sites by antibodies 7H9 gesting，性激素結(jié)合球蛋白抗體7H9的識(shí)別位點(diǎn) and 16D5 are not located in close proximity to the Olinked 和16D5不位于靠近O型掛鉤 glycosylation sites [Table 3a]. 糖基化位點(diǎn)[表3A]。 Similar findings, where 類似的結(jié)果，其中 SHBG le