بررسی مقایسه‌ای چند روش آزمایشگاهی برای جداسازی اگزوزوم‌های شیر گاو

نویسندگان

دانشگاه الزهرا

چکیده
امروزه اگزوزوم­های شیر به جهت دردسترس بودن و کارایی بسیار در محصولات آرایشی- بهداشتی و همچنین به عنوان نانوحامل­های دارویی، بسیار مورد توجه محققین قرار گرفته است. از آنجایی که یافتن روشی ساده و کارآمد برای جداسازی این وزیکول­ها بسیار با­اهمیت است، در این پژوهش به برخی روش­های جداسازی اگزوزوم از شیر گاو مانند اولتراسانتریفیوژ، جداسازی با استفاده از پلیمر PEG و چند کیت تجاری داخلی پرداخته و مشخصه­یابی اگزوزوم­ها با استفاده از DLS و میکروسکوپ الکترونی انجام شده است.

در روش اولتراسانتریفیوژ، به عنوان رایج­ترین روش جداسازی اگزوزوم، تعداد ذرات مشاهده­شده در تصاویر میکروسکوپ الکترونی بسیار پایین ±۲ پارتیکل در هر تصویر با بزرگنمایی تقریبی x ۶۰۰۰۰) برآورد شد، درحالی که در تصاویر میکروسکوپی کیت اگزوسان، تعداد ذرات زیادی (۱۵۰±۳۰پارتیکل در هر تصویر با بزرگنمایی تقریبی x ۳۰۰۰۰۰) قابل مشاهده بود. در جداسازی با استفاده از PEG، میانگین قطر ذرات با تکنیک DLS، ۲۶۳ نانومتر و بیشتر از روش­های اولتراسانتریفیوژ، کیت اگزوسیب و کیت آنااگزو که قطر ذرات به ترتیب ۱۷۶، ۱۴۲ و ۱۲۳ نانومتر بود، مشاهده گردید. میانگین قطر ذرات در تصاویر میکروسکوپی کیت اگزوسان، ۱۰±۷۰-۳۰ نانومتر بود که نتایج DLS هم کوچک­بودن سایز ذرات جداسازی­شده را تأیید کرد. با توجه به تعداد زیاد ذرات ریز (nm ۳۰ ) در نتایج میکروسکوپی کیت اگزوسان، این احتمال می­رود که چنین ذراتی به وفور در شیر وجود دارند و ممکن است روش­های دیگر قادر به جداسازی این ذرات ریز نبوده­اند. در نهایت هر چند تمامی روش­های مطالعه شده، قابلیت جداسازی اگزوزوم از شیر را داشتند ولی برای مقایسه دقیق­تر و معرفی یک روش استاندارد برای جداسازی اگزوزوم از شیر گاو، لازم است مطالعات گسترده ­­تری صورت پذیرد.

کلیدواژه‌ها


عنوان مقاله English

Comparative study of several laboratory methods for isolating exosomes from bovine milk

نویسندگان English

Hanieh Ghandchi
Reihaneh Ramezani
Zahra Moosavinejad
Alzahra University
چکیده English

Recently, milk exosomes have attracted much attention from researchers due to their availability and efficiency in cosmetic products and also as drug delivery nanocarriers. Since it is very important to find a simple and efficient method to purify these vesicles, in this research some methods of exosome isolation from bovine milk such as ultracentrifugation, using PEG polymer and several commercial kits were discussed and characterized. Detection of exosomes has been done using DLS and electron microscopy.

In ultracentrifugation, as the most common method of exosome isolation, the number of particles in the electron microscope images was estimated to be very low (5 ± 2 particles), while in the microscopic images of the Exosun kit, a large number of exosomes (150 ± 30 particles) was visible. In PEG precipitation, the average diameter of the particles in DLS results was 263 nm and more than the ultracentrifugation, Exocib and Anaexo kit, where the diameter of the particles was 176 nm, 142 nm, and 123 nm, respectively. The average diameter of the particles in the microscopic images of the Exosun kit was 30-70±10 nm, and DLS results confirmed the small size of the isolated particles. Considering the large number of small particles ( 30nm) in the microscopic results of the exosun kit, other methods may not have been able to isolate these small particles. Finally, although all the studied methods were able to isolate exosome from milk, more extensive studies are necessary to make a more accurate comparison and to introduce a standard method for isolating exosome from bovine's milk.


کلیدواژه‌ها English

exosome
extracellular vesicles
milk
polymer precipitation
ultracentrifugation
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  • تاریخ دریافت 18 خرداد 1405
  • تاریخ انتشار 18 خرداد 1405