Plaque Assay (for Bacteriophages)

The steps in determination of amount of virus by agar suspension plaque assay. This virus is a kind of bacteriophages. Its host cell is a bacterium.

The plaque is the area (spot) in which one viral particle has initially infected a cell (here, a bacterium) and has replicated itself in the cell producing new viruses. The cell is usually lysed upon the release of these newly produced viruses. The new phages go nowhere, but infect the nearby bacteria. Through many rounds of this life cycle, the spot grows big enough to be seen by naked eyes. (In the video, it looks like a hole in the agar) This spot, therefore, represents one virus at the start of the experiment. By counting the plaque number, the amount of virus in original sample can be calculated.

(Note: The remarkable studies of bacteriophage growth were originally pioneered by Ellis and Delbruck 69 years ago.)

Please rate or make a comment on this video.

Please also, if interested, make a discussion and comment on my blog about phages.
http://www.u-sabai-d.com/Bblog/index.php

Cheers.



วิดีโอนี้อธิบายขั้นตอนสำหรับปฏิบัติการหาปริมาณไวรัสโดยการนับจำนวนการเกิดพลัค พลัคเป็นบริเวณที่ปรากฏการเปลี่ยนแปลงของเซลล์เนื่องจากการติดเชื้อไวรัสโดยลักษณะกา­รเปลี่ยนแปลงที่พบบ่อยคือเซลล์แตกและตายเนื่องจากไวรัสที่ติดเชื้อเข้าไปอาศัยเซลล์ใ­นการเพิ่มจำนวน เมื่อดูเปรียบเทียบกับเซลล์ที่อยู่บริเวณข้างๆจะเห็นว่าบริเวณที่เกิดพลัคในกรณีที่เ­ซลล์ตายจะเป็นวงใส ขนาดของวงจะโตขึ้นเรื่อยๆเมื่อเวลาผ่านไป อย่างไรก็ตามขนาดของวงจะหยุดขยายหรือช้าลงหลังจากแบคทีเรียอยู่ในระยะที่มีการเจริญล­ดลง ทั้งนี้เพราะเฟจจำลองเพิ่มจำนวนได้ดีในขณะที่แบคทีเรียกำลังเจริญแบ่งตัวดี

Category:

Education

Tags:

• pfu
• plaque
• assay
• agar
• suspension
• phage
• bacteriophages
• titer
• virus
• amount
• determination
• พลัค
• ไวรัส
• ปริมาณ

[Seronegative2006]

Thank you for the invitation to the project. Let me reply you here so that my audiences with similar questions will also understand me and be informed.

It's very interesting,indeed, but I cannot make or accept it in this period of time (for 6-8 months at least) because I am running some projects for my academic career.

[pinknonsense]

You've done a very good job here - well done. I haven't seen this before. Please get in contact. I can make some suggestions about improving it - it could be extremely helpful.

[Seronegative2006]

Thank you very much. It's very kind of you. I do have some idea about it but don't have a chance yet. Your advices are most welcome and very appreciated. Cheers.

[arnchick]

It was very informative, I liked it. You may wish to add a bit showing the need for a control e.g. a negative control with bacteria and no phage. This will show if the method is conducted with the right concentration of bacteria if a homogenous lawn of bacteria is produced.

[Seronegative2006]

Thank you very much. I appreciate that.

The very very high diluted sample of phages (sometimes, no phages at all)are apparently the negative control by themselves.

In fact, in the clip "checking the result" there was a plate (on the left) which contained almost completely lawn of bacteria (only 2-3 clear holes seen).

It also confirmed that the enough bacteria were used.

Cheers.

[FarhadTv]

U r very rite, I think this is a very good job and a very good way of helping those who need it. I certainly did need it and I have improved my knowledge by watching this video.

Thanksa lot

[phunphage]

Thank you, for sharing this information. It would be very helpful to those who are suffering from MRSA if you could provide information on how to acquire, isolate and prepare a phage treatment. A cook book version for the layman would be perfect. Would you consider a collaboration on such a project?