Laboratory Management and Quality Assurance

Name: Rusydiana binte Kusni

Subject Title: Laboratory Management and Quality Assurance

Topic: Organization or regulatory bodies that regulate the operations of local laboratories in Singapore

Being one of the 8 professional centres of Health Sciences Authority (HSA) 2, Centre for Transfusion Medicine (CTM) is responsible in regulation of the operations of the bloodbanking section of clinical laboratories. It provides clinical support for the blood banks in the various local hospitals 2. These include recommendations on the most appropriate use of blood products and components, antibody identification and blood grouping discrepancies 1. CTM also draws out guidelines for the proper management, usage and safety of blood products 1. By doing this, CTM is able to meet its objectives of accomplishing finest quality of blood products supplied, while maintaining service excellence 2.

1. Clinical Services. (2007) Retrieved July 30, 2008, from Health Sciences Authority website: http://www.hsa.gov.sg/publish/hsaportal/en/health_services/transfusion_medicine/clinical_services.html

2. About Centre for Transfusion Medicine. (2007) Retrieved July 30, 2008, from Health Sciences Authority website: http://www.hsa.gov.sg/publish/hsaportal/en/health_services/about_ctm.html

Name: Raihana Binte Zainalabidin

Subject Title: Laboratory Management and Quality Assurance

Topic: Organization or regulatory bodies that regulate the operations of clinical laboratories in Singapore

The NEA (National Environmental Agency)
- formed in 2002, focuses on implementation of environmental policies
- branches out into 3 separate divisions that together helps ensure quality environment for Singaporeans
- Environmental Protection Division carries out programmes to help monitor, reduce and prevent environmental pollution 1
- controls disposal of waste from clinical laboratories in hospitals
o colour-coded bags, yellow, purple and red, are used for disposal of biohazardous,cytotoxic and radioactive wastes respectively
o 2 companies: Cramoil Singapore Pte Ltd and SembCorp Environmental Management Pte Ltd operates enclosed trucks that facilitate transport of wastes to hospital waste incinerators 2

1. About NEA. (2002). Retrieved 28 July, 2008, from: http://app.nea.gov.sg/cms/htdocs/category_sub.asp?cid=2


2. Toxic Wastes Control. (2002). Retrieved 28 July, 2008, from: http://app.nea.gov.sg/cms/htdocs/article.asp?pid=1531#CONTROL_OF_BIOHAZARDOUS_WASTES

Hello everyone. Hope all of you are enjoying your SIP, even if you are separated from your friends. For the past 5 weeks, I have been attached to several sections in the clinical laboratory.

1st week: Hematology
2nd week: Hematology
3rd week: Blood Banking
4th week: Order entry and Urinalysis
5th week: Clinical Chemistry

For this post, I am going to talk about the Hematology section. There are several tests that are done in this section. Examples would be Full Blood Count (FBC), Erythrocytes Sedimentation Rate (ESR), Retic count and Malaria test.

Full Blood Count (FBC)

For patient’s samples that are requested to undergo FBC, bloods are to be collected in an EDTA tube. This is to prevent the blood from clotting, as it can produce inaccurate results. To check for any clots that may have occurred, we just use a normal applicator stick. If there is even a small clot, the clot will stick to the applicator stick. If a clot is detected, we will reject the sample and a new sample is requested. At the same time, we will check if the sample is sufficient to be run. If it is less than 1.5ml, the sample will also be rejected. After making sure that there is no clot, the sample will be spun for mixing for about 5 minutes. After that, we will arrange the tubes in a rack before putting it in the machine to be run. The machine, Sysmex XE-2100, will do the FBC and the results will be uploaded to the LIS. If the machine detects some abnormalities in the blood, it will ‘trigger’ another machine which will produce a blood slide for further observation by the medical technologist. This machine, Sysmex SP-1000i, is solely responsible for making blood smearing and staining. So, after the samples have been run through the first machine, the rack will automatically be directed to the 2nd machine. Only the samples that are required to produce a slide will be aspirate out by the machine.

XE-ALPHAN ( Sysmex XE-2100 & SP-100i)

Retrieve from: http://www.sysmex-ap.com/default.asp?pageid=110

After all the tubes have undergone both machines, the results will then have to be validated by the medical technologist. For those samples that have slides being processed, the medical technologists will have to observe the slide under the microscope to view any abnormalities before validating. Abnormalities include increase or decrease in the number of the different type of WBCs, presence of any reactive lymphocytes, etc. If there are abnormalities, a manual differential count is done and the results will be keyed into the LIS.

In scenario where there is not enough blood for the machine to make a slide, a manual smearing and staining is required. So, a blood smear is done manually on a glass slide using another glass slide as a slider. After that, a manual stain will be done where the slide is covered with Wright stain and buffer pH7.2 for 10 minutes. It is then washed using distilled water and left to dry for another 10 minutes. It is then ready for observation. Another alternative for staining is to use the Sysmex SP-1000i under its manual mode where the slide will be put inside the machine manually before staining is carried out.


Retic count

Another test that is carried out in this section is the Retic count. This is to find out the number of retics or reticulocytes present in the blood. Retics are immature red blood cells (RBCs) that have not matured yet. It takes generally one day for the retics to mature and become fully mature RBCs.

To perform this test, a single drop of patient’s blood is mixed with a drop of the retic stain. The retic stain used is actually methyl blue stain. It is then incubated for 15 minutes at 37°C. After that, it will then be manually smeared to a glass slide. The slide is then observed under the microscope where the number of retics is taken note. A retic can be differentiated from mature RBCs by the presence of dark blue granules in its cytoplasm.

Reticulocytes

Retrieve from: http://sg.wrs.yahoo.com/_ylt=A0S0zvgOt4lI0WgBfn4u4gt./SIG=11qi13bn9/EXP=1217071246/**http:/www.umm.edu/imagepages/1491.htm

Thus, a manual count will be carried out where the number or retic is being track down while counting 1000 red cells. The number of retics is reported as a percentage of the total red cells. The normal range of retics for a healthy individual is 0.5%-2%. If the percentage is lower, it indicates that the bone marrow is not producing a normal number of RBCs. It may be due to several reasons like lack of folic acid, iron or vitamin B12 in the diet. To confirm, further tests are needed to diagnose the specific cause. If the percentage is higher, it means that the bone marrow is producing more red cells in response to a blood loss or treatment of anemia.



Differential count

Besides doing the test required, I also learnt how to do a differential count using a DC counter. Before doing the test, I have to know how to differentiate the different types of WBCs like lymphocytes, neutrophils, basophils, monocyte and eosinophils.

Neutrophil
Retrieved from: http://sg.wrs.yahoo.com/_ylt=A0S0zu1YvYlIdBsA.iou4gt./SIG=129lakjtm/EXP=1217072856/**http:/www.technion.ac.il/~mdcourse/274203/lect9.html

Lymphocyte

Retrieved from: http://sg.wrs.yahoo.com/_ylt=A0S0zu5uvolI6fMAStEu4gt./SIG=126nhnksc/EXP=1217073134/**http:/www.niagaracc.suny.edu/val/lymphocytes.html

Basophil

Retrieved from: http://sg.wrs.yahoo.com/_ylt=A0S0zu6hvolI__MA.JIu4gt./SIG=11ue3sbaf/EXP=1217073185/**http:/www.carlalbert.edu/dwann/tissue.htm

Monocyte

Retrieved from: http://sg.wrs.yahoo.com/_ylt=A0S0zvnZvolIg2YBZyMu4gt./SIG=12l6hhp65/EXP=1217073241/**http:/www.montgomerycollege.edu/~wolexik/205_histology__page.htm

Eosinophil

Retrieved from: http://sg.wrs.yahoo.com/_ylt=A0S0zvj.volIwmgBc4wu4gt./SIG=12fm7tkp7/EXP=1217073278/**http:/cellbio.utmb.edu/microanatomy/blood/Question_1bl.htm

It is advisable to start counting the cells when the RBCs are just ‘touching’ each other and not stacked together. This makes the counting easier and more accurate. When the number of WBCs has reached 100 cells, the counter will sound a ring as an indicator. The numbers of cells for each type of WBCs are recorded as a percentage. Any increase or decrease of each type may indicate different illnesses or disease the patient is suffering. Further tests is needed to confirm the diagnosis.

With this, i hope all of you have learnt something from my attachment. Till next time friends..

Take care and don't forget to enjoy yourself!! =D

Name: Nur Sofieyana Bte M.D Ismaeil

Class: TG02

Rai's SIP.

For the past 4 weeks, im attached to a Cytogenetics Laboratory. Written below is what i have learnt so far, enjoy =)

In a nutshell, the Cytogenetics lab processes patients sample to have their chromosomes analysed so as to conclude if patient has abnormalities in their chromosomes or not.

Basically, what I have learnt in the 4 weeks of my attachment to the Cytogenetics laboratory can be summarised into 5 parts.

Why perform cytogenetics test?
Why that certain sample is collected?
Summary of processes
Journey of my blood
Identification and karyotyping of chromosomes

Why perform cytogenetics test?

The few reasons include prenatal diagnosis where parents would want to find out if the baby in the womb is abnormal, diagnosis of neonates with abnormalities, diagnosing children with dysmorphic features/ developmental delay and couples who have frequent miscarriages who might want to know if they have any chromosomal abnormalities and patients with haematological malignant diseases.

Why that certain sample is collected?

For adults, peripheral blood is withdrawn from patient and used as a sample.
However, for pregnant women who wants to have a prenatal diagnosis, different type of sample is withdrawn for women of different gestation week due to several reasons.
For pregnant women between 10-12 weeks gestation, the sample ‘’chorionic villi’’ is extracted. U can know what is the ‘’chorionic villi’’ from the diagram below.

image taken from http://www.nwabr.org/studentbiotech/winners/studentwork/2007/WB_BA_TRONGTHAM/piccvs.jpg

Why this is extracted is because during this time, the chorionic villi are still new and small and would not cause serious bleeding to the mother when extracted. Also, the amniotic fluid cannot be withdrawn at this stage because the amniotic fluid is too little and if withdrawn, might cause dehydration to the fetus.

For pregnant women between 16-20 weeks gestation, amniocentesis is carried out. This is the withdrawing of amniotic fluid. At this stage, the amount of aminotic fluid is sufficient and this is confirmed by ultrasound.

For pregnant women of 22weeks gestation, fetal cord blood is extracted because by this time, the baby in the womb is already huge and taking up a lot of space in the womb, making it difficult to extract the amniotic fluid as there is a high risk of injuring the baby with the syringe.

Also, cord blood takes up to 7 days to obtain results, which is considered a short period of time compared to other samples, it is important that the lab technicians does not make a mistake in handling the sample tat might lead to wasting the sample because abortion can only be done up to 24th week gestation. This is especially critical for cases where the fetus is found to be abnormal and parents would want to consider aborting the child.
Here, we can see the importance of handling the samples with care, accuracy and precision. That is why the cytogenetics laboratory is a highly critical laboratory which could not afford to make mistakes and waste their samples and so, I am only allowed to observe the processes or try out the processes once or twice in cases where the samples are abundant.

Summary of processes

For each sample that comes into the laboratory, it will go through a common procedure that is

Cultured with cultured and colcemid is added. Colcemid is a mitotic inhibitor which causes the cells not to divide at metaphase. This is so that when the cells are analysed later on, the chromosomes would be seen and can be analysed.
Centrifugation and removal of growth medium. Changing of medium and cells are resuspend.
Cells are checked under microscope to check for growth. If sufficient amount of colonies are present, cells are ready for ‘’Harvest’’. If overcrowding os observed, cells are ‘tiffed’ (redistributed) and checked a few days later for growth.
Harvesting. Cells treated with hypotonic solution which increases the cell volume to allow the chromosomes to spread.
Cells fixed with fixative that draws out water out of cells so as to preserve them.
Cells transferred onto glass slide and allowed to dry.
Cells stained with Trypsin and Giemsa stain.
Slide is viewed under the microscope.
Using a microscope that is connected to the computer, an image of the desired metaphase can be captured and shown on the computer.
A specific programme is used to analyse the chromosome, karyotype, and analysed for any chromosomal abonormalities.

Journey of my Blood sample

Tuesday, 8th July
4pm:
Peripheral blood was withdrawn from my right arm and collected into a sodium heparin tube (anti-coagulant).

Blood set-up was done. This procedure includes centrifuging my blood sample for 5 minutes so that the buffy coat layer can be obtained. This is shown in the diagram below.

image taken from http://www.nsbri.org/HumanPhysSpace/index.html

The buffy coat layer is extracted from the tube and reconstituted into 2 separate tubes containing M199 media. 5 microliters of ‘PHA’ is added into each tube to stimulate the division of my T-lymphocytes present in the buffy coat layer.

Thursday 10th July
4pm:
50 microliters of MTX was added into each tube to synchronise the cells.

Friday 11th July
10am: 50 microliters of Thymidine added into each tube.

12pm: 50 microliters of Colcemid added to tube 1.

1.40pm: 50 microliters of Colcemid added into tube 2

*Colcemid, as mentioned earlier will allow chromosomes to be seen under the microscope later on. Exposure to colcemid for 2 hours will give rise to many metaphases with short chromosome fragments. Exposure to colcemid for 20 minutes will give rise to few metaphases with long chromosomes.

4pm: Harvesting. Addition of hypotonic solution and subsequently, fixative.

Monday 13th July

10am: Cells prepared on glass slide, stained and ready to be analysed using the computer system.

Below is the pictures of my chromosomes in a metaphase.

(u can click on the image to enlarge it, if u want to see my chromosomes =))

Then, I identified the chromosomes and place them in their respective groups, this is called "karyotyping" shown below. It is then analysed for any chromosomal abnormalities such as translocation, deletion and others.

It looks like my chromosomes are normal and I have no chromosomal abnormalities, Thank God. =) (I was a little nervous to analyse them)

Identification and Karyotyping

So how did I learn to identify the chromosomes one by one? =) That was really something. It couldn’t be learnt overnight and I had nightmares about chromosomes during the first week.
Altogether, it took me 3 weeks to practice everyday identifying chromosomes until I could get them all correct although sometimes there is still human error =).
During the 3 weeks, when I was struggling really hard to recognize the chromosomes, I prepared for myself a table which I could refer to during identification of chromosomes which can be seen below.

A little tedious, but hey, no pain no gain. I am happy now that I could identify chromosomes and I’m proud to say I’ve learnt something.

Hope all of u like my post.
Byebye. Take care everyone.

RUSY'S SIP

Hey all of you out there!!! I hope you guys are doing great at your respective attachment sites. As for me, I’m attached to Centre for Transfusion Medicine (CTM). And I can tell you guys that I am having a great time at the blood bank.

For the first three weeks, I have been attached to the cross-match and red cell reference (serology) labs. And now, I shall share with you one of the important tests that I had observed.

Subject Title: Blood Banking
Name of Test/Topic: Antibody Screening

In the cross-match lab, AbSC is done on almost every new blood sample that arrives. This is because there may be samples from previous patients with a record of medical history. For these people, AbSC is done only if the previous screening was done more that three days from the current date.

For example: First sample of patient arrives on 1^st July (Tuesday), and AbSC is done. The next sample arrives on 3^rd July (Thursday). AbSC is not done for this sample, as the time period for this sample from the previous one is only two days. Another sample arrives on 5^th July (Saturday) and this sample is then allowed to run an AbSC.

The reagents needed for AbSC are SPO (Singapore Panel O) cells, and plasma/serum of the patient. The SPO cells are prepared in-house by one of the senior lab officers. In CTM, three panels are used, compared to the two panels that we use in school.

In this lab, antibody screening is automated with the help of a machine called Techno TwinStation. Blood sample is to be centrifuged upon arrival to separate the plasma from the cells. The samples are then loaded into the machine where antibody screening will automatically commence. The machine uses the principle of AHG phase AbSC (antibody sceening) as it uses gel cards, which contain coombs’ reagent. The incubation of the reagents is also done in the machine. As soon as the process is over, the machine will produce the results on the touch-screen attached to the machine.

In the serology lab, AbSC is done manually. There are two phases of AbSC: saline phase and AHG phase.

For saline phase, cells and serum are mixed in a tube in a ratio of 1:2 (1 drop SPO cells, 2 drops patient serum). The reagents are allowed to react in room temperature for 30minutes, before spinning them at 3000rpm for 15seconds. This phase is for detection of cold antibody, mainly IgM.

For AHG phase, the gel cards are also used. However, the reagents are manually pipette into the wells of the card, where sensitization occurs during a 15minute incubation at 37^oC. The ratio of cells to serum when using the gel card however is reversed 2:1 (50ml SPO cells to 25ml patient’s serum). This is because weak antigen may not be detected if there is a higher amount of cells than serum in the gel card. After incubation, the cards will be spin down at the speed of 1000rpm for 10minutes. RBC agglutinates will be filtered through the column of gel according to size. A negative reaction will show that all the red cells are settled to the bottom of the column, while positive reactions will produce cells within the gel. This phase is for detection of warm antibody, mainly IgG.

After antibody screening is complete, antibody identification can be done to identify presence of specific antibody.

Techno TwinStation

Retrieved from: http://www.diamed.com/product_detail.aspx?id=848&navvis=

Coombs Anti-IgG gel cards

Retrieved from: http://www.diamed.com/product_detail.aspx?id=82&navvis=

I hope that all of you readers have gained some knowledge from my entry. Just shoot any comments and I will try my best to reply as soon as possible.

taking cares
peace out

Name: Rusydiana binte Kusni

Class: Tg 02

AmiR's SIP

Helllo my fellow med techs!! I hope you guys are having a great time during your attachments. Im in diagnostic bacteriology.. cool rite!! Hahaha for the 1st month I am in central processing area where the specimens are being processed. For this week, im assigned to do blood culture. It is a good experience as we get to go through the various types of microbiology cultures.

Blood culture is basically to test for any presence of microorganisms in the blood of the patients, usually fungal and bacteria infections. The blood taken from the patients is inserted into vials which will enter the BACTEC machine for incubation. What is a BACTEC machine??

Subject title: Blood culture
Topic : BACTEC machine

•BACTEC culture vials
•-Blue top is for aerobic bacteria culture
•-Yellow top is for anaerobic bacteria culture
•-Red top is for fungi culture
•-Sliver top is for blood culture for paedritic

What goes on inside the machine???

1) Presence of microorganisms causes metabolism of nutrients in the culture medium releasing CO2 into the medium
2) The CO2 produced will react with a dye in the vial sensor
3) Light Emitting Diodes (LEDs) modulated by the dye will illuminate the racks, activating the vial’s fluorescent sensors.
4) Instrument’s photo detectors take the readings of the fluorescence which corresponds to the amount of CO2 released by the organisms.
5) Raw data from detector is sent to rack microprocessor
6) Positivity analysis is performed in the microprocessor
7) Positive vial lamp illuminates, audible alarm sounds and positive station displayed on the monitor.

What happen when there is a postive result?

· When positive vials are identified, the lab technologist pulls them out of the incubation for further investigations.
· After the blood culture is flagged positive, a 6-fold dilution is made, with 0.2ml (8 drops) of blood to every 1 ml of saline, then subculture on appropriate plates
· For aerobic culture, we use blood agar plate(BAP) and maconkey agar plate(MAC)
· For anaerobic culture, we use BAP, MAC and anaerobic blood agar plate(ANA)
· The plates are incubated overnight.
· The culture is dispensed onto a glass slide and direct gram staining is done, using crystal violet, Gram’s iodine, acetone and safranin to see presence of gram positive cocci or gram negative bacilli.

Therefore, a positive result means that there is a bacterial or fungal infection in the patient’s bloodstream thus, needs to be treated immediately and a negative result means that there is a probability that a patient is suffering from an infection from other causes. Some microorganisms like viruses are more difficult to grow in culture and more testing may be required.

Okay now roughly i hope u guys understand what i am talking about. If you want to enquire about anything can just call me aites!! I guess this is where I say goodbye!! N i am awaiting for my next adventure this week!!!

Enjoy your SIP people!!!!!

Name: Mohamed Amir

Class: TG02