2、染色菌的選擇 

菌jun齡ling對dui革ge蘭lan氏shi染ran色se結jie果guo的de影ying響xiang是shi十shi分fen大da的de。我wo們men染ran色se過guo程cheng中zhong選xuan擇ze的de菌jun應ying該gai是shi處chu於yu活huo躍yue生sheng長chang期qi，這zhe樣yang的de細xi菌jun活huo性xing強qiang，生sheng理li特te征zheng明ming顯xian，所suo以yi染ran色se的de結jie果guo準zhun確que度du高gao
，一yi般ban情qing況kuang下xia不bu會hui出chu現xian誤wu差cha。培pei養yang時shi間jian較jiao長chang的de革ge蘭lan氏shi陽yang性xing菌jun
，由you於yu細xi胞bao老lao化hua，甚shen至zhi有you部bu分fen菌jun在zai染ran色se之zhi前qian就jiu已yi經jing死si亡wang或huo者zhe自zi行xing溶rong解jie了le，造zao成cheng細xi胞bao壁bi通tong透tou性xing增zeng加jia
，就jiu會hui呈cheng現xian出chu假jia陰yin性xing。以yi金jin黃huang色se葡pu萄tao球qiu菌jun為wei例li，金jin葡pu是shi典dian型xing的de革ge蘭lan氏shi陽yang性xing菌jun
，有you人ren曾zeng經jing統tong計ji過guo培pei養yang至zhi不bu同tong時shi間jian的de金jin葡pu的de革ge蘭lan氏shi染ran色se結jie果guo，結jie果guo表biao明ming，在zai金jin葡pu活huo躍yue期qi的de22h-26h之間，染色結果的準確率基本可以達到100%
，而超過26h之後
，準確率就開始下降，培養至36h時
，準確率大概會下降30%左右。所以我們在染色時，一定要選擇處於活躍期
、活性較強的菌落，在檢測過程中一定要嚴格的在國標要求的時間段內進行染色試驗，不能提前或者延後。

3、塗菌的狀態

在zai染ran色se最zui初chu的de塗tu布bu中zhong
，在zai挑tiao取qu菌jun體ti後hou應ying該gai在zai無wu菌jun水shui上shang塗tu成cheng薄bo薄bo的de菌jun膜mo。而er在zai塗tu布bu過guo程cheng中zhong，若ruo是shi菌jun體ti堆dui積ji，也ye會hui極ji大da影ying響xiang染ran色se結jie果guo的de準zhun確que性xing。菌jun落luo堆dui積ji過guo多duo，往wang往wang菌jun體ti之zhi間jian變bian得de毫hao無wu縫feng隙xi，而er其qi細xi胞bao壁bi的de成cheng分fen影ying響xiang造zao成cheng了le脫tuo色se的de情qing況kuang不bu佳jia，容rong易yi產chan生sheng假jia陽yang性xing的de結jie果guo。同tong時shi，在zai初chu染ran、媒mei染ran及ji複fu染ran的de過guo程cheng中zhong，應ying將jiang染ran液ye覆fu蓋gai整zheng個ge菌jun膜mo，避bi免mian一yi部bu分fen菌jun染ran色se而er另ling一yi部bu分fen菌jun沒mei有you染ran色se。因yin此ci在zai塗tu片pian過guo程cheng一yi定ding要yao將jiang菌jun膜mo塗tu得de少shao而er均jun勻yun，這zhe樣yang才cai能neng達da到dao最zui佳jia效xiao果guo。

4、媒染時間

meiranshijianduigelanshiyinxingjunderansejieguoyouhendayingxiang。youyumeiranshijianguochang，jiejingzizaidianyechangshijianzuoyongxiaguofenlaogujiehezaijuntixibaobishang，yingxianglejiujingdetuosexiaoguo

，congerhuidaozhizhezhongjiayangxingdexiaoguo。yidianxingdegelanshiyinxingjundachangaixishijunweili
，yourenzuoguotongji，meiranshijianyangekongzhizai40s-60s之內
，染色結果準確率基本可達到100%，而超過60s準確率會有一定的降低
，若媒染超過100s，準確率甚至可降低接近20%左右。因此在媒染過程中一定要注意媒染時間，控製在50-60s為宜。

5
、酒精脫色

jiujingtuoseyeshiyigeduijieguoyingxiangjiaodadecaozuoguocheng。gelanshiyinxingjunhegelanshiyangxingjunzuizhuyaodechayishixibaobitaijutangcenghouduhejiegou，gelanshiyinxingjunxibaobitaijutangcenghenbo，zhiduotanghanlianggaoyincizaijiujingtuoseshi，xijunxibaobideduotangchengfenhuibeijiujingrongjie
，zengdalexibaobidetongtouxing，jiejingzijidianfuhewujiuhenkuaibeijiujingxituo
，zhihoujiuhuibeishahuangfuranchenghongse

；而er陽yang性xing菌jun肽tai聚ju糖tang層ceng厚hou，脂zhi多duo糖tang少shao，酒jiu精jing隻zhi能neng起qi到dao脫tuo水shui效xiao果guo而er無wu法fa進jin入ru，這zhe樣yang結jie晶jing紫zi和he複fu合he物wu就jiu無wu法fa洗xi脫tuo出chu來lai使shi細xi胞bao呈cheng紫zi色se。因yin此ci，酒jiu精jing脫tuo色se時shi間jian短duan，脫tuo色se效xiao果guo不bu佳jia，會hui導dao致zhi陰yin性xing菌jun菌jun體ti內nei的de結jie晶jing紫zi和he複fu合he物wu不bu能neng有you效xiao的de全quan部bu洗xi脫tuo出chu來lai，就jiu會hui出chu現xian明ming顯xian的de誤wu差cha，使shi陰yin性xing菌jun出chu現xian假jia陽yang性xing。而er洗xi脫tuo時shi間jian過guo長chang，也ye會hui導dao致zhi陽yang性xing菌jun的de細xi胞bao壁bi被bei酒jiu精jing破po壞huai，導dao致zhi細xi胞bao內nei的de紫zi色se被bei洗xi脫tuo
，呈cheng現xian假jia陰yin性xing。因yin此ci洗xi脫tuo時shi間jian也ye是shi革ge蘭lan氏shi染ran色se的de關guan鍵jian環huan節jie。洗xi脫tuo時shi間jian應ying當dang嚴yan格ge控kong製zhi在zai20s-30s之間，同時保證洗脫效果。