ELISA Test for Virus Detection

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24/07/18 Sciences Reference this

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  • Zohrah Miakheyl

Immunology practical

Introduction

Viruses are small living organisms that requires living host for its survival and can lead to infection if it fully invades the host cells taking over the cell machinery to produce virus. There are many different viral infections including Measles, Mumps, Rubella, and Cytomegalovirus. Measles is an infection of the respiratory system caused by the measles virus. The measles virus is very contagious and can easily spread via sneezing, coughing and personal contact. Measles is known by the red-brownish rash starting on the face and spreading to the rest of the body. Other symptoms of measles virus include fever, diarrhoea, runny nose, conjunctivitis and dry cough (Permar et al, 2006). Mumps is an infection caused by the paramyxovirus. This viral infection is also highly contagious like Measles and spreads also via sneezing, coughing and direct contact. Mumps usually causes swollen salivary glands, fever, dry mouth and difficulties in swallowing. Rubella is an infection caused by Rubella virus, member of the genus Rubivirus. The spread of the virus is similar to measles and Mumps. Most common symptoms of this infection are a pink, light red rash lasting up to three days, mild fever, cold cough, sore throat, conjunctivitis and joint pain in adults (Stock, 2012). In contrast to Measles, Mumps and Rubella, Cytomegalovirus (CMV) usually don’t have any symptoms and therefore most people being infected may not know that they have CMV. People who are being infected often develop mild symptoms similar to flu or glandular fever. This viral infection can become serious in babies infected before birth, because this virus can be transmitted from mother to foetus. Cytomegalovirus is a virus belonging to herpes family of viruses. This virus is spread via body fluids (saliva and urine) and personal contact (Griffiths, 2002). The above stated viral infections usually occur at childhood, but it can occur at any age. Measles, Mumps and Rubella are often protected by giving the children MMR vaccination at 13 months and a booster is often given to three to five years old children. To find out from what the four patients are suffering from ELISA technique was applied to detect antiviral antibodies in patient’s serum for the viral infections Measles, Mumps, Rubella and Cytomegalovirus.

Method  

Results

Figure 1 microtitre plate showing the results of the four patients tested.

The ELISA technique shows that patient 1 is positive for Measles (IgM) and CMV (IgG), patient 2 is positive for Measles and Rubella (IgG), Patient 3 is positive for measles (IgG) and patient 4 is positive for CMV (IgM) and Measles, Mumps and Rubella (IgG).

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Discussion

Four patients were tested for viral infections using ELISA technique and the results showed positive for some patients for some of the viral infections. The ELISA results obtained were good and no errors were made. This might be due to accurate practical skills such as pipetting, appropriate filling of the wells, proper emptying and washing of the plate. Furthermore accurate serum dilution and measurement has also added to the accuracy of the results. Controlling the experimental conditions such as incubation time has also lead to improved results. To obtain better results volume measurement, dilutions, experimental conditions can be improved further. The volume measurement can be improved by accurately use of Gilson pipettes by ensuring there is no air bubbles and the pipettes are fully calibrated. Experimental condition can be improved by carefully monitoring the incubation duration.

Patient’s case study

Patient 1

A 56 years old patient was admitted with dense reddish brown maculopapular rash, headache, low grade fever, spot rash, mild abdominal discomfort and diarrhoea, high temp, mild conjunctivas symptoms. Throat swab done by the GP didn’t reveal any severe symptoms, but was referred for antiviral antibodies screening examination. After performing antiviral antibody screening the results showed positive for measles (IgM) and CMV (IgG). IgM is made during immune response in the early stage of infection and is mainly found in lymph fluid and blood where it functions as neutralizer against harmful agents. Whereas IgG is made in the late stage of infection and stays in the body for a long period of time. Due to its long survival period in the body IgG is useful for passive immunization. IgG is mainly found in blood, lymph fluid, cerebrospinal fluid and peritoneal fluid and plays a vital role in humoral immune response by activating the complement system (Lima, 2012). The sign and symptoms of the patients is similar to measles therefore indicating that the patients might be suffering from measles and might be in the early stages of the infection due to IgM found in the serum sample and macolopapular rash is also the sign of the infection at early stages. The mild abdominal discomfort, diarrhoea, and cold-like symptoms also indicates that the patient might suffer from CMV infections, but since IgG is found in patient’s serum sample, the infection might also be the result of immunisation and prior infection. All the symptoms of this patient are also the symptoms of measles therefore this patient might suffer from measles in the early stage of it, because of the antibody IgM and the reddish macolopapular rash is also found in the early stage of the infection) . Currently there are no means of preventing the developed measles infection, but the symptoms can be reduced using approved medications such fever-reducers, antibiotics, and vitamin A supplements (Permar et al, 2006).

Patient 2

A healthy 8 years old boy was examined by GP in a routine check and was referred for full blood counts. From the physical examination, the GP found that the boy was having faint cracking-like sound due to sever cold that he has been suffering since 3 weeks. Eczema-like rash was also observed, which was treatable by E45 creams, but the cause of this symptom was not clearly known as he has no family history of eczema. After his full blood count revealed no abnormalities and the cause of eczema was not known, his GP referred him for antiviral antibodies screening. The screening result showed that the boy was positive for Measles and Rubella (IgG) infections. The signs and symptoms of this patient appears to be different from the sign and symptoms and Rubella and Measles apart from the cold that the patient was suffering from. One of the first signs of Measles and Rubella infection is red-pinkish rash, where in Measles it appear to be observable for a week and in Rubella it lost for at least 3-7 days, but varies from patient to patient. The patient always had eczema-like rash, which means that the patient might not be infected by Measles or Rubella. The positive results obtained from antiviral screen might be due to immunisation and prior infections. Since the patient is not suffering from virus infection, the doctor ca refer him to Dermatologist for eczema examination. Since he is suffering from cold, the patient can take cold reducing medications (Gen,1999).

Patient 3

A 26 years old healthy female attend her GP for confirmation of her pregnancy test after she tested at home using home pregnancy test about a week ago and got positive results. She believes she is about 28 days pregnant, but this is her fist confirmation although she has been pregnant in the past and had early miscarriage about 10 months ago. GP preformed second pregnancy test and the positive results confirmed that she is pregnant. Since she was only 4 years old when moved to UK from Tajikistan, her medical record is available from the age of 6. Due to incomplete medical records, the patient is advised to carry out blood test so that her vaccination status can be known, therefore antiviral antibody screen was carried out and the results appear positive for Measles infection. The development of MMR vaccination appear to be very successful against this viral infection and due to improve and advancement of MMR vaccination, Measles infection has become less common now, but it shows to be dangerous among pregnant women if occur. This is because Measles might lead to miscarriage, stillbirth or early delivery if the pregnant individual is not immune against the infection. Although patient 3 do not display any sign and symptoms, but the antiviral antibody screen shows that she is positive for this infection. This positive result suggests that the miscarriage that she had might be due to Measles infection. However this cannot be said with certainty, because of her incomplete medical history. The antiviral positive result might be due to early infection or immunization. Since her medical history is not fully known, she probably needs MMR vaccination to make sure she is immunized against the viral infection

Normally the signs and symptoms of Measles infection tend to appear between 6 and 21 days. . Therefore it’s important that she is aware of any signs symptoms such as fever, cold and any skin rash that might appear at later stages of Measles infection. If she is infected then human normal immunoglobin (HNIG) treatment can be carried out to reduce the symptoms of infection. However this does not mean that miscarriage can be prevented (Permar, 2006).

Patient 4

A 43 years old male who had kidney transplantation 9 months ago due to kidney failure, which was caused as a result of Type 1 Diabetes Mellitus, which was diagnosed at the age of 11. The patient had a good diet and controlled blood glucose, but due change in life style and alcohol consumption and irregular insulin administration his blood glucose level has changed and become unpredictable. This life style has led to impaired kidney functioning, which resulted in estimated Glomerular Filtration Rate (eGFR) of 30ml/min. After monitoring eGFR about 3 months, a steady reduction was observed until the age of 39 with eGFR of 10ml/min. Although the patient was under dialysis, he was introduced to ambulatory peritoneal dialysis (CAPD) while waiting for suitable kidney donor for transplantation. After a few months CAPD treatment, he was referred to haemodialysis, which was carried out 3 times a week, because he could not get on with CAPD. After successful single kidney transplant, the patient shows to recover well from the surgery and the kidney appeared to function well. Following the checkups, the patient appeared to feel better and showed good recovery. However, the patient complained of generally unwell feelings with severe cold/ flu, where he has suffer for at least couple of weeks. Chest pain, abdominal cramps with diarrhea was also reported on this visit and physical examination shows some abdominal tenderness with no swelling or any other tenderness and normal blood pressure. The Auscultation examination however showed crackling and wheezing sounds. Microbial culturing was performed on stool and mid-stream urine samples a land with full blood count, creatinine measurement and an antiviral antibody screen examination.

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The microbial culture from urine samples revealed nothing significant and eGFR shows to be similar to last check up (10ml/min). The full blood count however shows that the Haemoglobin concentration was 12g/do, which is lower than the normal range (normal range 13-18g/dL) and this is thought to be as a result of impaired glomerular filtration rate, which is caused due to kidney failure and his haemoglobin concentration might be also low due to his kidney-transplantation and the immunosuppressive medication that he receives which causes mild bone-marrow suppression causing low hemoglobin concentration (Joist et al, 2006) . All other parameter however appear to be within the normal ranges indicating nothing significant. But the white blood count appears to be slightly higher than the normal range. The normal white blood count is 4-11×10 to the 9/L, whereas the patient’s count is 11.1×10 to the 9/L. This indicates that the patient might be suffering from viral infection. This is evident from the antiviral antibody screen examination, where the patient appeared to be positive for CMV (IgM), Measles, Mumps, and Rubella (IgG). Therefore it is the Measles infection that causes the while blood cell counts to alter. In general viral infections disrupts the bone marrow function temporarily and this leads to variation in white blood cell counts. The severe cold/flu, abdominal discomfort and diarrhea that the patient is suffering from is also the signs of CMV infection, this is because individuals who undergo organ transplantation are more susceptible to CMV infection (Rafailidis et al, 2012).The reason that the individuals are susceptible is because the medication they received reduces the immune system response to avoid rejection of kidney transplant (Cordero et al, 2012). However the patient does not display any symptoms for Rubella, Measles and Mumps, but the positive antiviral screening result might be due to previous infection or immunization. His crackling and wheezing sound might be result of cardiovascular disease that he might suffer from because of his lifestyle Kataoka, 2008).

In order to stay healthy the patient needs to improve his lifestyle by improving his diet by reducing/stopping alcohol consumption. If the symptoms of CMV becomes worse than antiviral drug can be used. If the immune system is temporary weakened then there is no need for any antiviral drug, because the patient will recover from the infection when the immune system is recovered.

List of references

Cordero, E. et al. (2012). Cytomegalovirus disease in kidney transplant recipients: incidence, clinical profile, and risk factors. Elsevier. [Online] 44 (3) p. 694-700. Available from: http://www.ncbi.nlm.nih.gov/pubmed/22483471. [Accessed: 15/02/2015]

Gen. (1999). Immunoglobulin (IgG) and (IgM) Antibody Responses to Rabies Vaccine .Lister Institute of Preventive Medicine, Elstree, Hertfordshire, U.K. 40 (10), p595-604.

Griffiths, P. D . (2002). Cause and Treatment Of Cytomegalovirus Infection.Journal of Antimicrobial Therapy. 49 (2), 243-255

Joist, H. Brennan, D.C. & Coyne, D.W. (2006). Anaemia in the kidney-transplant patient. Journal of The American Society of Nephrology. [Online] 13(1) p. 4-10. Available from: http://www.ncbi.nlm.nih.gov/pubmed/16412965. [Accessed: 11/03/2015].

Kataoka, H. & Matsuno, O. (2008). Age-Related Pulmonary Crackles (Rales) in Asymptomatic Cardiovascular Patients. Annals of Family Medicine. [Online] 6(3) p.239-245. Available from: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2384982/. [Accessed: 15/03/2015]

Lima, R. C. et al (2012). Interpretation of the presence of IgM and IgG antibodies in a rapid test for dengue: analysis of dengue antibody prevalence in Fortaleza City in the 20th year of the epidemic. Journal of the Brazilian Society of Tropical Medicine. [Online]. 45(2) p. 163-167. Available from: http://www.ncbi.nlm.nih.gov/pubmed/22534985. [Accessed: 22/03/2015].

Permar, S. R, Griffin, D. E and Letvin, N. L. (2006). Immune Containment and Consequences of Measles Virus Infection in Healthy and Immunocompromised Individuals.CLINICAL AND VACCINE IMMUNOLOGY,. 13 (4), 437–443.

Rafailidis et al,. (2012). Sever cytomegalovirus infection in apparently immunocompetent patients: a systematic review. Virology Journal. [Online] 5 (47) p. 1-7. Available from: http://www.virologyj.com/content/5/1/47 [Accessed: 11/02/2015]

Tingle, A. J, Allen, M, Petty, R. E, Ketthyles, G. D and Chantler, J. K. (2015). Rubella Associated Arthritis: Comparative Study of Joint Manifestation Associated With Natural Rubella Infection.Annals Review of the Rheumatic Diseases. 45 (1), 110-119.

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