HELLO PEOPLE! QINGLING'S BACK! :D
This month, I'm posted to the Thalassaemia section of the lab! The one that Felicia was at before me! I've learnt to perform five tests there and surprisingly it was pretty easy! My colleagues in the same lab as me were awesome! They taught me every test patiently despite me making mistakes along the way! They even left the test for me to perform all by myself whenever the test was ordered! So before i begin, im sure all of us know what a sickle cell looks like right? yes. like this!
Picture taken from
Therefore, in this entry, I’m going to share with you the Sickling Test! It provides a rapid screening test for Hb-S status of an individual and serves as a confirmatory test for the Alkaline Cellulose Acetate (the one that Felicia mentioned in one of the previous post) and Acid Gel electrophoresis. One important characteristic
of this test is that the sickling phenomenon occurs only with low oxygen tension. It will detect if the RBC will change into a sickle shape after mixing with the chemical that will
reduce the amount of oxygen carrying it.
A positive and negative control should be used in this test.
The method is as follows.
First, we have to prepare the reagent, which is a 0.2g of Sodium Metabisulphite. It is measured accurately using the weighing balance. After which, dissolve with 10mL of deionised water and mix it up and down using a pipette. 5 drops of the reagent is then added to 1 drop of anti-coagulated blood on the slide and mixed well. Immediately cover it with a cover-glass and leave for it to dry completely as the mixture may be present outside the cover glass. Seal the sides of the cover-glass with petroleum jelly-parrafin
wax mixture. Ensure that it covers the sides of the cover-glass properly as no oxygen should enter. Incubate at 37C for 2 hours. Finally, examine under microscope.
Sickling test should be negative in a normal individual. However in patients with Hb-S disease,
a marked sickling is usually visible.
One important thing to note while performing this test is that patients on medication of drugs such as phenothiazines dapsone will give a false negative result. Also old reagents or if preparation
is improperly sealed, will also result in false negative result.
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Isn't it simple? :D
If you have any doubts, dont hesitate to ask me!
Hello!
ReplyDeleteMay I know who's posting? (:
Anyway, I would like to know what is the use of adding Sodium Metabisulphite?
Thanks!
Siew Ming
0702862D
TG 02 Grp2
Hey Qing,
ReplyDeleteWhy is it that certain drugs will give a false negative? Does the drug affect the RBC or does it interfere with the reagent of the test?
Yvonee
0703189A
Hi!
ReplyDeletecan i ask, what is acid gel electrophoresis? (it's different from the Hb electrophoresis right?) are there any factors that can give false positive results?
LIM JIA HUI JOEY
tgo1 0703605F group 2
To SiewMing : Oops sorry! This is Qingling posting! Anyway sodium metabisulphite acts as a catala to hasten the reaction in the condition where there is low oxygen tension in order for the reaction to occur.
ReplyDeleteTo Yvonee : The drugs will affect the shape of the RBC thus causing false negative results when the RBC is supposed to look sickle in shape.
To Joey : The Hb electrophoresis is used as the initial procedure to screen for haemoglobin variants while acid gel electrophoresis is used to separate and confirm the haemoglobin variants that co-migrate in the Hb electrophoresis. Factors such as presence of Hb-C as well as when the preparation dries out will cause a false positive result.
Let me know if you guys still dont understand my explanation! :D
Oh.. I understand now.
ReplyDeleteThanks!
Siewming (:
Hi Qingling,
ReplyDeleteI would like to know why petroleum jelly-parrafin wax mixture is used to seal the sides of the cover glass. Any specific reason?
zi shuang
Qingling,
ReplyDeleteYou mentioned that the Sickling Test is only screening test for Hb-S status of an individual, so what are the additional confirmatory tests?
And regarding to the "Hb-S status", does it mean whether they have one/two gene? As I have read that people who have one gene — sickle cell trait — have a fairly small percentage of hemoglobin S. People with two genes — sickle cell disease — have a much larger percentage of the defective hemoglobin.
Li Yinliang Alex
TG02 0704894E
Group 8
19 August 2009
To Zi Shuang : Petroleum jelly-paraffin wax mixture does not oxidize on exposure to the air thus does not allow oxygen to enter when used.
ReplyDeleteTo Alex : The sickling test itself is the confirmatory test already while screening tests for HbS are the Alkaline Cellulose Acetate and Acid Gel electrophoresis.
Its not one/two gene. Its hetero and homozygous. This test detects both heterozygous and homozygous forms of sickling. Hb-S trait (heterozygous) has about 35% of Hb-S while Hb-SS (homozygous) is mainly about more than 90%.