【正文】 134 2 2 259 259 8 8 69 69 1 1 120 120 12 12 250 250 1 1 Control serum 對(duì)照血清 40 40 1 1 80 80 4 4 160 160 10 10 43 43 10 10 103 103 14 14 203 203 24 24 42 42 2 2 79 79 1 1 154 154 2 2 SHBG was assayed by ELISA using 11F11 as the coating antibody with the detection antibody as indicated. SHBG是通過檢測(cè)抗體酶聯(lián)免疫吸附使用11F11作為涂層抗體檢測(cè)。C. 在20 176。 / Steroids 64 (1999) 259–265 /類固醇64（1999）259265 Page 3 第3頁(yè) 100 l of SHBG standard plasma or patient plasma (1:1000 SHBG 100升的標(biāo)準(zhǔn)血漿或患者血漿（1:1000 dilution in assay buffer) was added to each well for over 緩沖液稀釋）以上，每孔加入 night incubation at 4176。 ELISA plates coated with 酶標(biāo)板涂有 polyclonal SHBG antibody and the addition of standards 0 to 400 nmol/l. 性激素結(jié)合球蛋白多克隆抗體和0到400 nmol / L的標(biāo)準(zhǔn)除了 Polyclonal antibodybound SHBG sequentially probed with monoclonal 多克隆抗體綁定SHBG順序探討用單克隆 antibodies, as indicated, with wash steps between each antibody. 抗體，如前所述，每個(gè)抗體之間的洗滌步驟 。 Partially purified 部分純化 nondeglycosylated human SHBG, lanes 2 and 4, probed with antibodies 非deglycosylated人類SHBG，2和4車道，探討與抗體 11F11 and 7H9 as indicated. 11F11和7H9表示。 (a) SDSPAGE Western blotting of human pregnancy plasma, lane （一）SDS PAGE電泳免疫印跡人類妊娠血漿，車道 1, and human male plasma, lane 2, probed with antibody 11F11. 1，人類男性的血漿，2巷，探討抗體11F11 。C with 簡(jiǎn)而言之，包被酶標(biāo)板4℃，過夜 intact rabbit antiSHBG serum diluted in PBS (29 l in 10 完整的兔抗SHBG血清稀釋在PBS（29日升 10 ml PBS) 100 l per well. 毫升PBS）中，每孔100升。 Using of a Behring 使用一個(gè)貝林 ELISA ProcessorII (Hoechst, Germany) allowed semiau ELISA處理器II（德國(guó)赫司特，）允許半AU tomation of processes. 進(jìn)程tomation的。C, after which the plates were washed, 緩沖區(qū)）在20小時(shí)1℃，分別洗凈后，板， and substrate solution was added. 和底物溶液加入。C. 在4 176。 Phos 磷 phatebuffered saline was M NaH phate， M的NaH 2 2 PO 寶 4 4 , M NaCl, ， M氯化鈉（NaCl）， adjusted to pH with 5 M NaOH. 。 Beckton Dick 酶標(biāo)板（獵鷹3912微量三。 Immunization and cell fusion 免疫和細(xì)胞融合 Female RBFDN mice were immunized with 10 g of 女RBF DN小鼠免疫10克 SHBG in plete Freunds adjuvant at 4week intervals. 在完成Freunds輔助SHBG每隔4周。 Email address: johnL2 (JG Lewis) E mail地址：johnL2（JG劉易斯） Steroids 64 (1999) 259–265 類固醇64（1999）259265 0039128X/99/$ – see front matter 169。 * Corresponding author. *通訊作者。 Materials 材料 Human SHBG was obtained from Scripps Laboratories, 人類性激素結(jié)合球蛋白是從斯克里普斯實(shí)驗(yàn)室獲得， San Diego, CA, USA and Antimouse IgG1peroxidase and 加利福尼亞州圣迭戈，美國(guó)和Antimouse IgG1的過氧化物酶和 antimouse IgG2aperoxidase from Nordic Immunology, Pil antimouse IgG2a北歐免疫過氧化物酶，弼 berg, Netherlands. 伯格，荷蘭。 In 在 addition, we report their use in a sameday, simple, and 此外，我們?cè)谕惶欤?jiǎn)單報(bào)告其使用，并 direct enzymelinked immunosorbent assay (ELISA) for 直接酶聯(lián)免疫吸附試驗(yàn)（ELISA） human SHBG in plasma. 人類血漿中SHBG。 It is apparent that low levels of 很明顯，水平低 SHBG may be associated with conditions of excessive an SHBG條件可能與過度的一個(gè) drogen action [2], and there is recent interest in the rela 氫行動(dòng)2]，且有在近期利益關(guān)系 tionship between SHBG and plasma lipoproteins [3]. 性激素結(jié)合球蛋白和血漿脂蛋白之間tionship [3]。 Keywords: Monoclonal antibodies。 The monoclonal antibodybased ELISA shows 單克隆抗體為基礎(chǔ)的ELISA顯示 excellent performance characteristics and is unaffected by added testosterone or estradiol. 性能優(yōu)良的特點(diǎn)，是增加睪丸激素或雌激素的影響 。 Some在同一天有些 of the antibodies were selected to form a basis of a sameday, nonpetitive, enzymelinked immunosorbent assay (ELISA) for SHBG 被選中的抗體在HSBG的基礎(chǔ)上在血漿中，進(jìn)行非競(jìng)爭(zhēng)，酶聯(lián)免疫吸附試驗(yàn)（ELISA） in plasma.。 accepted 23 November 1998 1998年5月22日，1998年11月 23日 Abstract 摘要 Four monoclonal antibodies to human sex hormonebinding globulin were raised and characterized. 提出了四個(gè)人類性激素結(jié)合球蛋白的單克隆抗體和特點(diǎn) 。Page 1 第1頁(yè) Monoclonal antibodies to human sex hormonebinding globulin 人類性激素結(jié)合球蛋白的單克隆抗體 (SHBG): Characterization and use in a simple enzymelinked （SHBG）：表征在一個(gè)簡(jiǎn)單的酶聯(lián) immunosorbent assay (ELISA) of SHBG in plasma 性激素結(jié)合球蛋白（ELISA）檢測(cè)血漿中的免疫吸附試驗(yàn) John G. Lewis*, Nicholas J. Longley, Peter A. Elder 約翰G 四分之三Three of the four antibodies四分之三個(gè)抗體 recognised different antigenic determinants on SHBG. 公認(rèn)的SHBG不同的抗原決定簇 。 The assay employs a purified IgG2a SHBG monoclonal antibody adsorbed to the wells of a microtitre plate. 該試劑盒采用純化IgG2a性激素結(jié)合球蛋白單克隆抗體吸附到一個(gè)微孔板井。 169。 Sex hormonebinding globulin。 In 在 addition, the role of unliganded plasma steroidbinding pro 此外，unliganded血漿中的作用的類固醇結(jié)合蛋白 teins bound to membrane receptors as a delivery mechanism 作為一個(gè)傳遞機(jī)制的約束膜受體蛋白 of steroids to target cells has attracted much attention [4,5] 靶細(xì)胞的類固醇備受關(guān)注[4,5] and established a functional link to steroid hormone recep 并建立了一個(gè)功能鏈接到類固醇激素受體 tors [6]. 職權(quán)范圍[6]。 Although others have reported 雖然其他人已經(jīng)報(bào)告 immunoradiometric and ELISA assays for SHBG [7,8], this SHBG [7,8]，放射免疫和酶聯(lián)免疫吸附檢測(cè) is also the first report of an ELISA using exclusively mono 也是第一份報(bào)告，使用專門的單聲道的 ELISA clonal antibodies. 單克隆抗體。 Antimouse Igperoxidase and isotyping Antimouse免疫球蛋白過氧化物酶和isotyping kits were from Life Sciences, Amersham, UK. 從生命科學(xué)學(xué)院，英國(guó)Amersham公司，試劑盒。 PO Box 151, Christchurch, New Zealand. 郵政信箱151，新西蘭基督城。 1999 Elsevier Science Inc. All rights reserved. 0039128X/99 / $ 見前面問題169。 One week after the third injection, spleens were excised and 一個(gè)星期后的第三次注射，脾臟切除 spleen cells fused with FOXNY myeloma cells at a ratio of FOX NY骨髓瘤細(xì)胞融合的比例脾細(xì)胞 5:1 as previously described [9]. 5:1如前所述[9]。 Beckton迪克 inson, Oxnard, CA) were coated overnight at 4176。 The following day, the 次日， plates were washed four times with PBS containing % ％的四倍 Tween20 (v/v) and blocked with assay buffer [PBS con 吐溫20（V / V）和阻止緩沖液PBS CON taining % Tween20 (v/v) and % gelatin (w/v)] 150 ％的Tween 20（V / V）％明膠（W / V）] 150 l/well for 5–10 min at 20176。 C The following day, the plates were again washed, 翌日，板塊再次洗凈， and 50 l of assay buffer was added to each well, followed 緩沖液和50升，每孔加入，其次 by 50 l of supernatant for a further 2h incubation at 20176。 The substrate was freshly 基板是新鮮 prepared by the addition of 40 mg of o phenylenediamine 編制由40毫克 鄰苯二胺 and 60 l of 30% H 60升30％的H 2 2 O 216。 . 。 The following day the plates were 翌日板