WHAT IS A SERUM PROFILE?

Serum profiles are based on the detection of circulating antibodies. This can be done by using any of the  techniques now available, in order to obtain the necessary information about the immunological status of an animal at that specific moment or in previous stages. It is slowly becoming more common to use these studies in the sanitary control of pig farms. They are, in many cases, an important tool for reducing expenses and increasing the sanitary level. 

Serological studies allow one to, among other things: know the sanitary status, choose the best time for vaccination, control vaccination programs and avoid the introduction of new diseases to the farm.  

We must take the following factors into account in order to perform a good serologic study of a farm: 

What do I wish to know anout my farm? What is the question I want to ask? Can serology give me an answer to my doubts?
2.   About which animals should I ask these questions and when?
How many samples do I have to take and how should I go about doing it?
What are the differences beween the different laboratory techniques?

1. What do I wish to know anout my farm?
   This would be the first question we must answer before starting any kind of study. What do we want to know about our farm? As we will see later, serology can help us to know certain aspects, but it is of no use in other things. It is very important to know these limitations of  serology. When planning any serological study we must take into account, among other things: the dynamics of the appearance of the antibodies, the difficulties of discerning between disease and vaccine-induced antibodies, the time needed for antibodies to be at detectable levels, etc.  

Do I want to know if a disease is present or not on my farm? How is the vaccination program working? What is the sanitary status of the newly arrived animals..? These are some of the questions that can be asked. When explaining the applications of serology we will see how to get these studies under way.   

2. About which animals should I ask these questions and when?
   This is another important question we must define. We must remember that animals can have different serum profiles at different stages of their productive life. Therefore we must sample those animals that will provide the answers. For example, we must take into account that immunoglobulins can not go through the placental barrier, so a piglet will not have immunoglobulins until it has received the maternal colostrum. 

From the moment the piglet sucks colostrum it will passively acquire humoral immunity. This immunity will be similar to the mother's. The maximum intake takes place between 6 to 36 hours after birth, and we can detect immunoglobulins in the bloodstream between 12 and 24 hours after colostrum ingestion. The length of time these antibodies remain in the blood stream is variable and will depend on the type of immunoglobulin, the mother's immunological status, the infectious agent, etc. Usually colostrum immunoglobulins can be found in weeks 3, 8 and 20. This is one reason among others as to why it is important to know at what age we should take the sample from the animal and from which age group it should be taken, i.e.; sows, piglets, etc.

3. How many samples do I have to take and how should I go about doing it? 
   This is the main question that has to be answered if we want the results to be representative of the actual situation. Besides the sampling from different areas of the farm and from different production areas, it is also important to determine how many samples are necessary to take from each area. There are two different aspects that must be evaluated on a porcine farm: those aimed at determining the presence or absence of a disease and, in the positive cases, those aimed at discovering the prevalence of the disease. In other words, the questions would be: Is this disease present on my farm? In the case that it is, how prevalent is it ? To answer these questions it is of the utmost importance to know the size of the sampling we need. This size will depend on the degree of accuracy we want to obtain. There are some tables (see other  sources of information at the end of the chapter) that indicate the number of samples needed depending on the confidence level we want to have, and the suspected prevalence, the precision and the level of confidence. The number of samples that must be taken varies depending on the desired level of confidence and on the suspected prevalence. In general, the following figures can be applied when performing any serological sampling:    
   
   Sows:

• Farms with less than 25 animals: every animal must be studied. 

• Farms with 25-100 sows: 25 samples from different animals that will be rotated in the following samplings. 

• Farms with more than 100 animals: 30 samples from different animals that will be rotated in the following samplings. 

In a feeding herd:

• At least 30 samples for each feeding area. 

The other question would be: what kind of sample must be taken and how? Serum profiles are mainly performed using serum, even though some times it is also interesting knowing the levels of immunoglobulins in milk and colostrum. Colostrum samples must be taken within 24 hours after parturition. If the sample is going to be immediately sent to the laboratory it must be kept refrigerated. If the sample is going to be shipped in the following days, it must be kept frozen.

Blood used for serum collection is usually obtained from: 

Ear marginal vein. Using a lancet and collecting the blood in a tube or using syringes with 16 or 25 mm gauge needles. This system has the advantage of being easy to perform, but the amount of blood collected is small, and contaminated samples are frequent. 

Tail vein. Usually with syringes (25 mm gauge needles), vacutainers or partial amputation. This method is also easy to perform, but the sample size obtained is only about 0.5 ml. It is not the ideal method. 

Jugular vein. Is one of the most frequently used methods, especially in sows. Usually with syringes, vacutainers, or monovet. The amount obtained is about 10 to 30 ml. 

Cava vein. Used to bleed all type of animals, from piglets to adults, and specially when a large amount of blood is needed. Needles vary depending on the size of the animal (10 Kg: 25 mm; 45 Kg: 38 mm; 100 Kg: 50 mm; sows of 100 Kg: 100 mm)

And finally, what do I do with the blood after the extraction? Blood must be kept at room temperature (in a cool place) until it coagulates (from one to two hours). After this, it must be kept at 4ºC overnight. Then, it is recommended that the blood be centrifuged, or at least, separate the serum from the coagula before shipping it to the laboratory. Blood tubes must be properly labeled and wrapped. If the study requires two different determinations with an interval of 21 days (serconversion) it is better to freeze the serum and wait until the next sample is taken in order to send both samples at the same time. A good diagnostic depends largely, on a good sample. Avoiding contaminations or serum alterations is a guarantee of a good diagnosis. 

4. What are the differences between different laboratory techniques?

As we mentioned at the beginning of this chapter, there are several laboratory techniques which are available nowadays  for serological studies. The diagnostic capability of a technique is determined by the evaluation of its sensitivity and specificity compared to the reference technique. Some concepts must not be forgotten when making the assessment of the available techniques. These are:

Sensitivity:  Is the capability of a method to detect positive sera. Sensitivity allows the detection of all positive cases.

Specificity: Is the method's capability to discriminate between positive and negative sera. A good specificity must lack false positive cases. No negative case must be considered as positive by any technique.

Predictive value: Is the capability of the technique to discriminate between animals that are suffering a given disease and animals which are not. In other words, it predicts the sensitivity and specificity for either a negative or a positive case. The predictive value can be: 

Positive: It is the frequency of disease among animals with a positive result.

Negative: It is the frequency of the absence of disease among animals with negative results.

Efficacy: Percentage of correctly classified animals.

True positive: Sick animal that has been correctly classified by the technique. 

False positive: Animal that has not been correctly classified by the technique.

The assessment of the sensitivity, specificity, and predictive value of any technique is performed by comparing its results with those obtained with a technique of reference or with the real disease, using the following formula: 

A: Positives for both techniques.

B: Real positives that are considered as negative by the assessed technique.

C: Positives of the assessed technique that are real negatives.

D: Negatives for both techniques.

Predictive value for positives:
A / A + C

Predictive value for negatives:
C / B + D

Among the techniques that are currently being used, those with better sensitivity and specificity levels are seroneutralization and ELISA. Depending on the laboratory, similar results can be obtained using both techniques.  The most important thing is to remember that if we want to compare results at different times, results at different stages... the same technique should be used, and even the same laboratory. If not, different results could be obtained and the diagnostic would not be correct.