201120445 六、發明說明： 【發明所屬之技術領域】 本發明係有關-種_食品巾動物賴朗的方法及檢驗試鑛組，特別 是以利用雜奈米粒子之雜標記免_定配合_磁減量的方式來做檢 驗0 【先前技術】 動物用藥品及含藥物飼料添加物雖可維護畜禽水聽物健康，促進畜牧事 業發展，但若不當棚齡造成食品巾之朗，或可祕遞抗菌株， 引起食品安全之疑慮，直接或間接危害國民健康。 為預防並遏止動_藥品概祕絲騎，配合衛生署訂定之「食σ 動物用藥殘留標準」，藥檢局自92年起即協調縣市衛生局，稽杳傳統3 超級市場、生鮮超市、商號、肉鋪、餐廳、小吃店、海產店等，抽驗市 農畜禽水產品及其加工製品等進行動物用藥殘留檢測，檢測項目包括氯 素、經四環黴素、氯四環黴素、脫祕四環黴素、四環黴素、諾伯徽素 2卡巴精 '乙型受體素類、孔雀綠、還·孔雀綠、硝㈣代謝物及 咖卿，賴編Μ嶋物用_ So。/〜/Γ的不合格率相對較高，其中雞肉不合格率約舰，烏骨象 率二_%〜33 3% ’ _ 14 6% ’蜂產品中蜂王聚❼ :::3_。雞肉檢出卿 乃卡巴於二Γ主要為恩氟奎林賴’其次_氧_環黴素，雞内臟1 類抗‘。财4 saIbut_〗*喊，蜂王漿為氣黴素及四環《 、▲㈣水產1^巾勒祕職抽驗絲，自類n不A格 率相對較高，香$之不騎“ 木，、、、頰產口口的不合格 香'，、、之不„袼率為5.3%〜22.2%; •鳟魚為5.3%〜6娜，·石斑 201120445 魚為27.2% ;吳郭魚為％，瑕類為10·0%。其中蹲魚為檢出氣徽 素及硕基知代謝物風險高產品；香魚不符規定藥品項目為恩I奎林緩酸 及硝基呋喃代謝物；吳郭魚為氯黴素及還原型孔雀綠；鰻魚及石斑#為硝 基呋喃代謝物及還原型孔雀綠。 由上述資料來看，動物用藥前餘食品__甚高，而檢測必然會持續 進行，以保護國人的健康。目侧來檢測水產品 要是液相層析質譜儀（LC/MS/MS)，其對各種動物用藥殘留的檢驗靈敏度 約是零點幾個PPb (〜1G，。使用LOMS/MS進行動物_殘留，固然二 精確’但在商業細上，仍呈現出許多_ ;如必須對所取得的水產品^ 行繁雜且冗長㈣纽財後，才能制LC/Ms/Ms量_殘含=; LCMS/MS的操作’必須在2〇_25 〇c的環境下，因此需要一個特別的=間 來安置LC/MS/MS ; LC/MS/MS的精確操作與維護，實不容易，故需聘靖 :位受過_丨_人貞，專職貞㈣作鋪器，無形_ 了該技術^ 應用層面，亦增加檢測成本。 用it 研究團隊欲使用EL及其他既有之技術進行動物 樂殘留檢驗’然而常因檢測時的操作複雜度、或過高的偽陽偽陰性，而 …法將該種技術普遍地推廣，此’市場上仍期待能開翻—種在水產品 中動物用_留之檢測技術，以符合市場所需。 【發明内容】 ==_編_純嶋㈣觸，包括·多個磁性 201120445 活性劑層，披覆於該磁性核上，以及待測的動物用藥抗體’結合於該界面 活性劑層上。 本文中所指之「磁性核」，其材料包括但不限於Fe3〇4、FeA、麻咏、 C〇Fe2〇4及NiFe2〇4，於較佳的實施例中，磁性核的材料係&心4。 本文中所指之「界面活性劑」係指能使目標溶液表面張力顯著下降的物質， 鲁以及降低兩種液體之間表面張力的物質，界面活性劑—般為具有親水與疏 水基團的雜兩性分子，可溶於有機溶液和水溶液。於本發日种，所使用 之^面活性册料包含但不限於有機酸、蛋白質A、蛋白f g、葡聚醋或 •微脂粒’於較佳的實施例中，界面活性劑的材料為葡聚醋。 本發明中，可測試的動物用藥係包含但不限於氣黴素㈣㈣咖喊 CAP)、細環黴素、氣四環黴素、脫氧經四環徽素、四環徽素、諸伯徽素、 乃卡巴精、乙型受體素類、孔鶴、顧型孔雀綠、絲料代謝物及啥 φ = (Q olone)類於較佳之實施例中，本發明檢測之動物用藥為氣徽素。 於本文中’所指的崎係包含任何可食狀動物，於較麵實施例中，動 物為水產動物，於更佳之實施例中，此水產動物為蝦。 树明另提供—種檢測食品中動物_殘留的方法，包括：提供本發明之 萃取檢體内^留的動物用藥，將萃取物與試劑混合後，量測磁減量訊號， 比對又測藥物之磁減量訊號標準曲線，以定量動物贿的殘留量。 201120445 由於每顆單獨磁性核的磁偶極方向 顆磁性核的磁偶極方向都是指峨的方向^時，由於熱運動，每 的，因此，當檢戦劑無外加磁 方向的機率都是-樣 交流變化時，、， ’並不會表現出雜。當外加磁場為 而使試劑產生妹子因為磁作用，會隨外加交流磁場轉動，因 生化率。㈣試纖樣品混錢，部 抗動物用藥抗體會與剌樣品巾 m 、 積變大，P駭1 齡，糾雜奈米粒子體 待馳細 _觸絲+嶋。域-來，會使血 油結合之磁性奈餘子無法再隨料加紐磁場轉動，而造 會°^ ;因此，糊㈣她麵_物用藥， 夕的磁性奈米粒子與其結合，而使試劑的交流磁化率降低越多。因 2檢職品t磁化率減少多少的量，即可比對該動物_磁減量訊 唬私準曲線，以定量動物用藥的殘留量。 為配合本發檢驗方式，本發明另提供―種萃取檢體内__物用藥 的方法，其步驟為：⑴、將檢體打成泥狀，（2)、加人萃取液萃取後離心留 上清液，⑶、以低壓真雜發法(使用抽氣幫浦協助萃取液的揮發），使該 上清液濃縮麟，且(4)、以獅齡驗濃縮乾驗。本發财法中之萃 取液為有機溶劑，於較佳之實補中，萃取液為乙酸乙自旨；於較佳的實施 例中，稀釋液則為二甲基亞砜以及磷酸鹽緩衝液。若想要得到較佳的萃取 率，步驟(2)可重複多次以提高萃取率。此萃取方式萃取程序需時約⑼分 鐘，現行用於LC/MS/MS的萃取程序需時約⑽分鐘，大大的縮減了萃取 的時間，並且不需要再用正己烷進行繁雜的萃取步驟。本發明所開發出的 萃取程序’相較於現行用於LC/MS/MS的萃取程序，能有效提升萃取時效， 並增加萃取程序的使用便利性。且本萃取程序中所使用的儀器較為簡易。 201120445 例如現行祕LC/MS/MS的萃取伽1要賴細銳機(哪丽 evapom叶於本發明的萃取程序中，不需使用，而是以—個簡易幫浦來取 代。如此’不僅可降地檢峨本，亦有助於將來對祕用藥_檢測的推 廣。 本發明之檢測食品帽物賴前的方法，財之動物軸熱包括但不 限於氣黴素、触環黴素、氣四環黴素、脫輸四環黴素、四環黴素、諾 鎌素、乃卡巴精、乙鼓體義、孔絲、職魏錢、硝基咬喃代 謝物及啥諾剛Quinolone)類，較佳實施例之動物用藥為氯徵素。 【實施方式】 之内 下列實施罐代表本發明幾種獨祕及概]羽於_本 容0 實施例一試劑合成 將化學計量比率為1:2的硫酸亞鐵七水(叫7恥）與氣化鐵丄 (⑽卿)混合的鐵溶辦量的侧祕使其作為四氧化頌的 面活性劑樣使其分散在水中。該混合物加熱至Μ·9 驗： 定形成黑色的四氧化三鐵粒子。叢聚物以及多餘__以離心的^ 移除’並賤膠過朗析已舫高度濃_韻體。該 ' 的碌酸鹽緩衝液稀釋已獲得合適的濃度。 阳-7. 為了使抗聽合至錄奈綠子㈣㈣㈣上，叫鎌素抗晃 201120445 (Ab35658 - 500，Abeam)為例，加入NaI〇4溶液至磁性溶液中，使葡聚醣 氧化以得到醛基(-CHO) ’之後’葡聚醣可透過^取队的連結與抗氯黴素抗 體反應，因而能使抗氯黴素抗體共價的連接到葡聚醣上，如圖_ω所示。 未接合的抗氣黴素抗體則可透過磁性分離，從溶液中移除，此試劑可於檢 體内的待測樣品結合’而造成磁化率的降低，如圖一(b)所示。 使用磁力顯微鏡(magnetic force microscope，MFM)，量測到所合成出的氣 黴素檢測試劑中的磁性粒子粒徑分佈如圖二所示。粒徑平均值在纪励，標 準差為3.5 nm。 為確定磁蹄子表®有接合上驗素碰，本實_伽—端財fitc榮 光的二級抗體，來標示披覆在磁粒子表面的氣黴素抗體，如圖三(a)所示。 實驗中’加入過量的FITC ;級抗體與披覆有氣黴素抗體的磁性粒子混合。 充分混合後，再藉由磁性分離技術，將混合液中的磁性粒子分離出，而未 與磁性粒子細之驗素抗體接合的FITC二級紐，料具有雖，會留 在原混合液中。將分離出的磁性粒子再分散到pBS溶液中。由於螢光標記 FITC可吸收495 run波長的紫外光，並發散528肺波長的綠光，因此，將 此溶液放人#細微鏡下觀察，首先會制綠色亮點，如肛⑼巾箭頭所 指之處。此免點是FITC所發出的光亮。繼之，在溶液旁擺上磁鐵。此時溶 液中的磁性粒子會受到磁鐵的吸引而移動。偶若磁性粒子表面已接上氣徽 素抗體’附有FITC的二級抗體會應與磁絲子上的錄素抗舰合，而披 覆在磁珠表面上。當磁珠受職_則而移麟，圖三⑼所觀察到的綠 色壳點，也會往磁鐵的方向移動。本實施例中，將磁鐵放於圖三(的的下方， 磁性粒子應雜下方觸，因此圖三綱綠色亮點也麟往下方移動。此 201120445 預測在圖三(C)中被觀察到，這證實磁性粒子上，確實已接上氣黴素抗體 (Anti-CAP) ° 實施例二氣擻素含量的磁減量標準曲線建立 不同量的氯黴素(C1863-25Q Sigma)溶於pH-7.4的填酸鹽緩衝液以獲得不 同濃度由O.lng/g至30ng/g的標準氣黴素溶液。每種濃度的標準氣黴素溶 液作三重複的磁減量(IMR)測試，測試方法如下：40 μΐ的磁試劑與60 μ1的 不同濃度的標準氣黴素溶液於玻璃管中混合，IMR檢測使用的磁試劑濃度 為 0.1 emu/g (= 5 mg-Fe/ml)。先以磁性免疫分析儀(Xacpr0-El〇l，MagQu)量 測未結合成氯黴素-抗氯黴素抗體-葡聚醣-奈米磁性粒子前的XaeQ交流信 號’之後，不同濃度的混合物於室溫下形成氯黴素-抗氯黴素抗體-葡聚醣_ 奈米磁性粒子，量測並紀錄其交流信號χ^,φ，透過量測出的知,。及χ^φ，可用 下列的公式計算出IMR訊號。 IMR (%) = (χΜ，。- χΜ)/χκ,。X 100 % 對每一個給定濃度的樣品溶液，其時間依賴之Xac交流信號皆檢測三重複。 φ 可獲得如圖四之IMR訊號為氣黴素濃度((J)Cap)函數之實驗圖。圖四中可見 IMR訊號隨著氯黴素濃度的增加而降低，而當磁性免疫分析儀(XacPro-ElOl， MagQu)分析的IMR訊號幾近雜訊(約為虛線所示之0.82 %)時，氯黴素的 濃度低於0.1 ng/g。 由以前的試驗結果可知，待測物濃度與IMR訊號符合下列之邏輯特函數 (Logistic Function) A-B IMR(%) =201120445 VI. Description of the invention: [Technical field to which the invention pertains] The present invention relates to a method and an inspection test group for a food towel animal, Lai Lang, in particular, a miscellaneous label using a hybrid particle Reduce the amount of the way to do the test 0 [Prior Art] Animal drugs and drug-containing feed additives can maintain the health of livestock and poultry, promote the development of animal husbandry, but if the age of the shed is caused by the food towel, or secret Resistance to strains, causing food safety concerns, directly or indirectly endangering national health. In order to prevent and curb the _ medicines and secrets, in line with the "Standards for Residues of Foods for Animals" set by the Department of Health, the Drug Inspection Bureau has coordinated the county and city health bureaus since 1992, and has traditionally supermarkets, fresh supermarkets, and trade names. , meat shops, restaurants, snack bars, seafood stores, etc., testing the city's agricultural and poultry aquatic products and processed products for animal drug residue testing, testing items including chlorine, tetracycline, chlorotetracycline, off Mitetracycline, tetracycline, Norbertin 2 kaba fine 'B-receptor class, malachite green, also · malachite green, nitrate (four) metabolites and coffee, 赖 Μ嶋 _ _ So . / / / Γ 不 相对 相对 相对 相对 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡 鸡The chicken was detected in the case of Nikab in the second sputum, which was mainly enflukalin. The second was _oxy-cyclomycin, and the chicken viscera was resistant. Cai 4 saIbut_〗 * shout, royal jelly for pneumomycin and four rings ", ▲ (four) aquatic products 1 ^ towel Le secret examination silk, self-class n not A rate is relatively high, incense $ does not ride "wood,,, The unqualified incense of the buccal mouth is ',,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,, The cockroach is 10·0%. Among them, squid is a high-risk product for the detection of gas ash and abundance of metabolites; the squid does not conform to the prescribed drug project as a meta- sulphuric acid and nitrofuran metabolite; Wu Guoyu is chloramphenicol and reduced malachite green ; squid and grouper # is a nitrofuran metabolite and a reduced type of malachite green. According to the above information, the pre-existing food for animal use is very high, and the test will continue to be carried out to protect the health of Chinese people. The side of the eye to detect aquatic products is liquid chromatography mass spectrometry (LC/MS/MS), which has a sensitivity of about a few PPb (~1G) for various animal drug residues. Animals_residues are performed using LOMS/MS. Although the second precision 'but in the fine business, there are still many _; if the aquatic products obtained must be complicated and lengthy (four) New Zealand, then the amount of LC / Ms / Ms can be made _ residual = = LCMS / MS The operation 'must be in the environment of 2〇_25 〇c, so a special = room is required to place the LC/MS/MS; the precise operation and maintenance of the LC/MS/MS is not easy, so it is necessary to hire Jing: The position has been _丨_人贞, full-time 贞 (4) as a paver, invisible _ the technology ^ application level, also increases the cost of testing. With IT research team wants to use EL and other existing technologies for animal music residue test 'but often Due to the complexity of the operation at the time of detection, or the excessive false negative of the pseudo-yang, the technology has been widely promoted, and this is still expected to be able to open up in the market - the detection technology for animals used in aquatic products In order to meet the needs of the market. [Invention content] ==_编_纯嶋(四)Touch, including ·Multiple magnetic 2011 20445 an active agent layer coated on the magnetic core, and the animal drug antibody to be tested 'bonded to the surfactant layer. The magnetic core referred to herein includes materials including but not limited to Fe3〇4, FeA, paralysis, C〇Fe2〇4 and NiFe2〇4, in a preferred embodiment, the magnetic core material & heart 4. The term "surfactant" as used herein refers to the surface of the target solution. A substance whose tension is significantly lowered, and a substance which lowers the surface tension between the two liquids. The surfactant is generally a hetero- and amphoteric molecule having a hydrophilic and a hydrophobic group, and is soluble in an organic solution and an aqueous solution. The surface active material used includes, but is not limited to, an organic acid, a protein A, a protein fg, a vinegar or a oligosaccharide. In a preferred embodiment, the material of the surfactant is vinegar. In the test, the animal drug system includes, but is not limited to, pneumomycin (4) (4) coffee CAP), fine ring mycin, gas tetracycline, deoxytetracycline, tetracycline, zhubo, and Kaba fine, beta-receptor, Kong He, Gu-type malachite green Silk material metabolites and what φ = (Q olone) class in the preferred embodiment, the present invention is of animal drugs for the gas detection element emblem. The sirloin referred to herein is any edible animal. In the more preferred embodiment, the animal is an aquatic animal. In a more preferred embodiment, the aquatic animal is a shrimp. Shuming also provides a method for detecting animal_residue in food, comprising: providing animal medicine for extracting the body in the invention, mixing the extract with the reagent, measuring the magnetic reduction signal, comparing and measuring the drug The magnetic decrement signal standard curve is used to quantify the residual amount of animal bribes. 201120445 Since the magnetic dipole direction of each magnetic core in the direction of the magnetic dipole is the direction of the ^, because of the thermal motion, each, therefore, the probability of the detection agent without the applied magnetic direction is - When the exchange changes, , ' does not show impurities. When the external magnetic field is applied, the reagent is generated by the girl due to the magnetic action, and will rotate with the external alternating magnetic field due to the biochemical rate. (4) The sample of the test fiber is mixed with money. The anti-animal drug antibody will be mixed with the sputum sample towel m, and the product will become large, P骇1 age, and the nanoparticle particles will be finely _touched + 嶋. Domain-to, will make the blood and oil combined with the magnetic Naizi can no longer rotate with the magnetic field of the material, and create a ^^; therefore, paste (four) her face _ substance medication, the magnetic nano-particles combined with it, so that The more the AC susceptibility of the reagent is reduced. Because the amount of magnetic susceptibility of the 2nd inspection product is reduced, the animal's _magnetic reduction signal can be compared to the residual amount of animal medication. In order to cooperate with the method of the present invention, the present invention further provides a method for extracting an in vivo __ drug, the steps of which are: (1), the sample is made into a mud, (2), and the extract is extracted and centrifuged to leave The supernatant, (3), using the low-pressure true hair method (using the pumping pump to assist the evaporation of the extract), the supernatant is concentrated, and (4), the lion age test concentration test. The extract in the present invention is an organic solvent. In the preferred embodiment, the extract is acetic acid. In a preferred embodiment, the diluent is dimethyl sulfoxide and a phosphate buffer. If a better extraction rate is desired, step (2) can be repeated multiple times to increase the extraction rate. This extraction method takes about (9) minutes, and the current extraction procedure for LC/MS/MS takes about (10) minutes, greatly reducing the extraction time and eliminating the need for complex extraction steps with n-hexane. The extraction procedure developed by the present invention can effectively improve extraction aging and increase the ease of use of the extraction procedure compared to current extraction procedures for LC/MS/MS. And the instrument used in this extraction procedure is relatively simple. 201120445 For example, the current secret LC/MS/MS extraction gamma depends on the fine sharp machine (which is not required to be used in the extraction procedure of the present invention, but is replaced by a simple pump. Checking the transcript will also help to promote the use of the secret drug in the future. The method for detecting the food cap of the present invention, the animal axis heat includes but is not limited to pneumomycin, cyclosporine, gas tetracyclic Taxomycin, tetracycline, tetracycline, norepine, naicarbamate, b-bubble, perforation, occupational Weiqian, nitro-methane metabolite and quinolone Quinolone The animal drug of the preferred embodiment is chlorin. [Embodiment] The following implementation tanks represent several unique secrets of the present invention and are summarized in the first embodiment. The reagents are synthesized to synthesize ferrous sulfate heptahydrate (called 7 shame) with a stoichiometric ratio of 1:2. The side effect of the mixed iron solution of the gasified iron shovel (10) is dispersed as water as a surfactant of ruthenium tetroxide. The mixture was heated to Μ·9 test: black iron oxide particles were formed. The plexus and the excess __ are removed by centrifugation and the sputum is too thick. The 'di-acid salt buffer dilution has been obtained at the appropriate concentration. Yang-7. In order to make the anti-hearing to the Nai Nai (4) (4) (4), as the example of the alizarin anti-shock 201120445 (Ab35658 - 500, Abeam), add the NaI〇4 solution to the magnetic solution to oxidize the glucan to obtain The aldehyde group (-CHO) 'after' glucan can react with the anti-chloramphenicol antibody through the linkage of the team, so that the anti-chloramphenicol antibody can be covalently linked to the glucan, as shown in Figure _ω Show. The unconjugated anti-pneuxin antibody can be magnetically separated and removed from the solution, and the reagent can be combined with the sample to be tested in the sample to cause a decrease in magnetic susceptibility, as shown in Fig. 1(b). Using a magnetic force microscope (MFM), the particle size distribution of the magnetic particles in the synthesized gastrin detection reagent was measured as shown in Fig. 2. The average particle size is in the excitation, and the standard deviation is 3.5 nm. In order to determine the magnetic hoof table®, there is a secondary antibody that binds to the surface of the magnetic particle, as shown in Figure 3(a). In the experiment, an excess of FITC was added; the graded antibody was mixed with magnetic particles coated with a gasmycin antibody. After thorough mixing, the magnetic particles in the mixed solution are separated by a magnetic separation technique, and the FITC secondary material which is not bonded to the magnetic particle fine test antibody is left in the original mixed solution. The separated magnetic particles were redispersed into the pBS solution. Since the fluorescent marker FITC can absorb 495 run wavelength of ultraviolet light and divergence 528 lung wavelength green light, so the solution is placed under the #microscopic observation, first green highlights, such as the anus (9) towel arrow At the office. This exemption is the light emitted by FITC. Next, place a magnet next to the solution. At this time, the magnetic particles in the solution are attracted by the magnet and move. Even if the surface of the magnetic particles is attached to the gas-enhanced antibody, the secondary antibody with FITC should be attached to the magnetic surface of the magnetic beads. When the magnetic beads are employed, then the green shell points observed in Figure 3 (9) will also move in the direction of the magnet. In this embodiment, the magnet is placed in the lower part of Figure 3, and the magnetic particles should be touched underneath, so the green highlights of Figure 3 are also moved downward. This 201120445 prediction is observed in Figure 3 (C). It was confirmed that the magnetic particles were indeed attached to the gas-mycin antibody (Anti-CAP) °. The magnetic reduction standard curve of the gas content of Example 2 was established to establish different amounts of chloramphenicol (C1863-25Q Sigma) dissolved in pH-7.4. The acid salt buffer is used to obtain standard gasmycin solutions with different concentrations ranging from 0.1 ng/g to 30 ng/g. The standard gasmycin solution of each concentration is subjected to three repeated magnetic reduction (IMR) tests. The test methods are as follows: 40 μΐ of magnetic reagent was mixed with 60 μl of different concentrations of standard gasomycin solution in a glass tube. The magnetic reagent used for IMR detection was 0.1 emu/g (= 5 mg-Fe/ml). The instrument (Xacpr0-El〇l, MagQu) measures the XaeQ AC signal before binding to chloramphenicol-anti-chloramphenicol antibody-dextran-nano magnetic particles, and the mixture of different concentrations forms at room temperature. Chloramphenicol-anti-chloramphenicol antibody-dextran _ nano magnetic particles, measuring and recording its exchange letter For the χ^, φ, the measured value of the transmission, and χ^φ, the IMR signal can be calculated by the following formula: IMR (%) = (χΜ, .- χΜ)/χκ, X 100 % for each For a given concentration of the sample solution, the time-dependent Xac AC signal is detected for three repetitions. φ The experimental image of the IMR signal as the gasmycin concentration ((J)Cap) is obtained as shown in Fig. 4. The IMR signal can be seen in Figure 4. As the concentration of chloramphenicol decreases, the concentration of chloramphenicol is low when the IMR signal analyzed by the magnetic immunoassay analyzer (XacPro-ElOl, MagQu) is nearly random (about 0.82% of the dotted line) At 0.1 ng/g. From the previous test results, the concentration of the analyte and the IMR signal meet the following Logistic Function AB IMR(%) =
B 201120445 其中，A ' B、φ。及β是該函數的參數。將實驗數據套入邏輯特函數中，可 得到Α=0.81、Β = 4.54、φ〇 = 60.4及β = 0.61，並以實線繪於圖四中。而實 驗數據與logistic fimction的相關係數r2= 0.98，顯示兩者有高度的相關性， 顯不DVIR-(j)cAP確為邏輯特函數。 實施例三氣黴素的IMR干擾測試 由於食品中不一定只含有一種動物用藥，因此需要試驗含有不同的動物用 藥，是否對IMR的檢測造成干擾。於此實施例中，使用了其他典型的動物 用藥，如孔雀石綠(MG)、還原型孔雀石綠(LMG)、3 -氣基-2 -嚼唑烷酮 (AOZ，富來頓之代謝物）、磺胺甲基嘧咬(SMR)、以及羥四環素(〇TC)作為 氯黴素之IMR干擾測試之用。此六種溶液之濃度及編號如下表—所示，並 添加了相㈤濃度的嫌雜吻於其t。除了職1之外，其·液的干擾 物質皆分別加人了 IGng/g至溶財。而後，仙抗驗素抗體試劑來測試 六種溶液的IMR訊號，絲如圖五所示。依據㈣辑顯示，溶液糾與 溶親間的户值是0.74、溶液#1與溶液約_户值是〇·23、溶韻與溶 液料間的ρ值是0.38、溶液#1與溶液#5帛的ρ值是〇 42、溶液糾盥溶液 桃間的减是〇.25。依據T test統計，ρ值大純％時，表示兩社間不 明顯差異。這麵溶祕之磁魅峨與其絲权_量域，彼此間 無顯著差異。耻’制韻量氣黴素糊檢職黴素含量時，不會受到 其他動物用藥的影響。 201120445B 201120445 where A ' B, φ. And β is a parameter of the function. Putting the experimental data into the logical function, you can get Α=0.81, Β=4.54, φ〇=60.4 and β=0.61, and draw them in Figure 4 in solid lines. The correlation coefficient between the experimental data and the logistic fimction is r2=0.98, which shows that there is a high correlation between the two, and it is obvious that DVIR-(j)cAP is a logical special function. Example III IMR Interference Test for Pneumomycin Since foods do not necessarily contain only one animal drug, it is necessary to test whether different animal drugs are used, and whether it interferes with the detection of IMR. In this example, other typical animal drugs such as malachite green (MG), reduced malachite green (LMG), and 3-alcohol-2 - chemozolidone (AOZ, Fulton's metabolism) were used. , sulfamethylpyrimidine (SMR), and hydroxytetracycline (〇TC) were used as IMR interference tests for chloramphenicol. The concentrations and numbers of the six solutions are shown in the table below, and the phase (f) concentration of the kiss is added to its t. In addition to job 1, the liquid interference substances are added to IGng/g to dissolve money. Then, the anti-testin antibody reagent was used to test the IMR signals of the six solutions, as shown in Figure 5. According to the (4) series, the value of the solution between the solution and the solution is 0.74, the solution #1 and the solution are about _ household value is 23 23, the ρ value between the solution and the solution is 0.38, solution #1 and solution #5 The ρ value of 帛 is 〇42, and the reduction between the solution remedy solution is 〇.25. According to the T test statistics, when the value of ρ is large, it means that there is no significant difference between the two communities. There is no significant difference between the magnetic enchanters and the silk weights. When the shame's rhythm is used, it will not be affected by other animal medications. 201120445
1 5 ng/g CAP 2 5ng/gCAP 及 10ng/gMG 3 5 ng/g CAP 及 10 ng/g LMG 4 5 ng/g CAP 及 10 ng/g AOZ 5 ------ 5 ng/g CAP 及 10 ng/g SMR 6 ~~—--— 5 ng/g CAP 及 10 ng/g OTC 實施例四IMR檢測動物用藥之萃取方式 圖六顯示此發明的檢測動物用藥之萃取方式。以蝦為例，去除蝦殼後，將 蝦肉打成泥狀(10分鐘），加入乙酸乙醋組成的萃取液6毫升與1〇克的瑕肉 混合’ 3分鐘後’離心5〇〇〇rpm，5分鐘，得到上層上清液，大部份的瑕肉 中氯黴素即於此上清液中。為了增加萃取率，此加入萃取液的步驟可重複 多次。此上清液以低壓真空揮發法使該上清液濃縮賴，乾燥物加入二甲 基亞職以及顧鹽緩衝液構成的稀釋液溶解，即為可直接上機檢測的樣 #品。此萃取方式萃取辦需_⑽分鐘，大编輯了萃取的時間。 為了檢視此萃取方式是否確可萃取出蝦肉中殘留的氯徵素，於瑕泥中加义 100 ng的氣黴素，以模擬含有10ppb氯黴素的瑕肉。經圖六的萃取程序後 使用紐伽_，断韻量跡4複触綠量_果為臆❶/ % °根據圖四_素檢_曲線，此磁減量訊號對顧 的濃度、核_pb，表示圖六_的萃取流程，萃取率可達約㈣。如增如 乙酸乙酯萃取步驟的程序，則可更加的提高萃取率。 201120445 【圖式簡單說明】 圖一、（a)本發明之檢測試劑示意圖；（b)檢測試劑接合待測物之示意圖。 圖二、合成出之氣黴素檢測試劑中的磁性粒子粒徑分佈。 圖二、表面接有氣黴素抗體(Anti-CAP)的磁性奈米粒子與附有FITC之二級 抗體的(a)示意圖及(b) (c)在螢光顯微鏡下觀察到受磁鐵吸引的移動 行為。 圖四、氣黴素磁減量檢測之標準曲線。 ^ 圖五、含有不同干擾物之溶液對氯黴素磁減量訊號之影響比較。 圖六、本發明實施例四之從蝦子中萃取氣黴素之流程圖。 【主要元件符號說明】 益 121 5 ng/g CAP 2 5 ng/g CAP and 10 ng/g MG 3 5 ng/g CAP and 10 ng/g LMG 4 5 ng/g CAP and 10 ng/g AOZ 5 ------ 5 ng/g CAP And 10 ng/g SMR 6 ~~---- 5 ng/g CAP and 10 ng/g OTC Example 4 IMR Detection Animal Extraction Method Figure 6 shows the extraction method of the invention for detecting animal drugs. Take shrimp as an example. After removing the shell of the shrimp, pour the shrimp into a mud (10 minutes), add 6 ml of the extract of ethyl acetate and mix with 1 gram of meat. After 3 minutes, centrifuge 5 times. The supernatant was obtained at rpm for 5 minutes, and chloramphenicol was the supernatant in most of the meat. In order to increase the extraction rate, the step of adding the extract may be repeated a plurality of times. The supernatant is concentrated by a low-pressure vacuum evaporation method, and the dried product is dissolved in a dilute solution of a dimethyl group and a salt buffer, which is a sample which can be directly detected by the machine. This extraction method takes _ (10) minutes to extract and edit the extraction time. In order to check whether the extraction method can extract the residual chlorophyll in the shrimp meat, 100 ng of gasmycin was added to the mud to simulate the meat containing 10 ppb of chloramphenicol. After the extraction procedure of Figure 6, use New Zealand _, the rhyme trace 4 re-touch the green amount _ fruit is 臆❶ / % ° according to Figure 4 _ prime test _ curve, the magnetic decrement signal to the concentration of the Gu, nuclear _pb , which indicates the extraction process of Figure 6 and the extraction rate can reach about (4). If the procedure of the ethyl acetate extraction step is increased, the extraction rate can be further increased. 201120445 [Simplified description of the drawings] Figure 1. (a) Schematic diagram of the detection reagent of the present invention; (b) Schematic diagram of the detection reagent bonding the analyte. Figure 2. Particle size distribution of magnetic particles in the synthesized gasmycin detection reagent. Figure 2. (a) Schematic diagram of (b) magnetic nanoparticles coated with anti-CAP and antibiotic-attached secondary antibody and (b) (c) attracted by magnet under fluorescence microscope The behavior of the move. Figure 4. Standard curve for the detection of pneumomycin magnetic decrement. ^ Figure 5. Comparison of the effects of solutions containing different interfering substances on the chloramphenicol magnetic depletion signal. Fig. 6 is a flow chart of extracting pneumomycin from shrimps according to the fourth embodiment of the present invention. [Main component symbol description] Benefit 12