1. Antigen Leukocyte Cellular Antibody Test (ALCAT)<br />Purpose of the Tool<br />The antigen leukocyte cellular antibody test (ALCAT) measures changes in leukocyte (white blood cell) size/volume by quantity following incubation with food and other test agents, such as food additives, dyes, environmental chemicals, pharmacoactive agents in foods, molds, antibiotics, and NSAID’s in vitro (Akmal, Khan, & Khan, 2009; Wüthrich, 2005). ALCAT has been developed to analyze minute cellular changes in leukocytes when they are exposed to foods to identify food sensitivities and intolerances (Brady, 2010). This tool claims methodology to detect responses on any potential pathway (IgG, IgA, IgM, IgE, IgD, complement, pharmacologic, toxic, lectin, etc.). Foreign entities in whole blood can cause autolysis, also known as autocytotoxicity, and other cellular reactions when interacting with antibodies within the blood. When collected blood samples are passed through an aperture, the cellular reactions are detected by a computerized device called the ROBOCat II, manufactured by Cell Science Systems, Ltd, (Akmal et al, 2009; Pasula, 1993).<br />ALCAT claims diagnostic capabilities in identifying substances responsible for sensitivity and intolerance reactions, particularly for delayed response allergic reactions. This test is being marketed directly to the public and health professionals as more effective than serum specific IgE tests or traditional skin prick tests. The conditions that are claimed to be identified by ALCAT testing include: migraine, headaches, ADD/ADHD, autism, insomnia, depression, anxiety, bed wetting, allergies, hay fever; asthma, post nasal drip, chronic sinusitis, irritable bowel syndrome (IBS), inflammatory bowel disease (Crohn's disease, ulcerative colitis), acne, eczema, psoriasis, urticaria, Candida, autoimmune diseases (Hashimoto's thyroiditis, rheumatoid arthritis, lupus, multiple sclerosis), fibromyalgia, metabolic syndrome, obesity, infertility , gastroesophageal reflux (GERD), poor memory, unexplained chronic fatigue, and weight loss (Allergy Society of South Africa, 2009).<br /> <br />Explanation to Patients<br />Given that most patients will not have much, or limited, education in biology or hematology, it is pertinent that the purpose and methods of ALCAT be explained in a clear and concise fashion. It would be described by stating that the patient’s blood will be drawn, then small amounts of that blood sample will be put into a solution with extracts from foods called “test agents.” These agents may react with certain cells in the blood through a pathway such as an immune response. The leukocytes (white blood cells) are responsible for making antibodies to fight bacteria, viruses, and other foreign matter. Proteins, dyes, and other constituents of food may trigger this immune response by the leukocytes; therefore, the “test agents” are representative of the proteins, dyes, and other food constituents (Akmal et al, 2009).<br />The blood is then exposed to different types of solutions that will burst the red blood cells, leaving behind only the leukocytes to be detected by the computerized device, ROBOCat II. The ROBOCat II uses electrical signals to detect any changes in the leukocyte size/volume; this information is then sent to a computer, which forms a graph of the sizes and the number of leukocytes (ALCAT, 2011). The graph, called a histogram, can be interpreted to reveal whether the patient has sensitivities or intolerances to any of the “test agent” foods and to which ones (Wüthrich, 2005).<br />History of the Tool<br />Antigen leukocyte cellular antibody test (ALCAT) is a blood test developed over 24 years ago to identify non-IgE-mediated hypersensitivities, intolerances, and adverse reactions to foods (ALCAT, 2011; Wüthrich, 2005). IgE and IgG antibodies related to certain food constituents may not be cause of all reactions. There is also much controversy regarding IgG testing, in addition to the theories that frequent exposure to a particular food may result in high levels of IgG as a protective biological response. Therefore, ALCAT was developed as an alternative assessment tool. ALCAT detects the positives that also appear on ELISA, but this tool has the advantage of finding more reactions that are occurring on pathways other than IgE and IgG (Brady, 2010).<br />Mechanism<br />The testing begins by an aliquot of citrated blood being diluted to a 1:5 dilution with buffer, 90μl is added to each test agent, which are diluted preparations of food extracts of standardized for potency (50% glycerol, 50% water with 1:10 dilution of antigens) following 45 minutes incubation at 37°C with agitation (Akmal et al, 2009; Neetling & Kachelhoffer, 1998). The samples are then incubated at room temperature for 45 minutes (Neetling & Kachelhoffer, 1998). Red blood cells are lysed by adding 16 mL Isotone II 0.5% alcalyse, an azide-free, electrolytic solution, supplied by Cell Science Systems, Ltd. The ROBOCat II then analyzes each test agent with one control from every 10 food items (Akmal et al, 2009). The white blood cells pass through a narrow aperture, and the number of cells and the size of each cell are determined by measuring changes in electrical resistance (Wüthrich, 2005; ALCAT, 2011). A histogram is generated: relative number of counts on the y-axis; and cell size (in femtoliters) on the x-axis (Neetling & Kachelhoffer, 1998).<br />A baseline distribution curve is compared to the distribution curve generated by the analysis of each test agent, resulting in a calculated difference between the curves and the standard deviation (SD). A non-reactive result (NEG) is any reactivity under SD1; marginally reactive (RANGE 1+) is reactivity between SD1 and SD2; reactive (RANGE 2+) is reactivity between SD2 and SD3; and markedly reactive (RANGE MPOS) is greater than or equal to SD3 (Akmal et al, 2009).<br />Safety of Use<br />The safety of ALCAT has not been thoroughly explored, but some studies state the safety of ALCAT if the patients’ or subjects’ symptoms did not worsen. No found studies have mentioned if symptoms worsened during the course of an elimination diet based on ALCAT results. It is important to note that sterile needles and appropriate sterilizing dressing be used during blood withdrawal for the testing (Neetling & Kachelhoffer, 1998). A false negative result is possible, as with any test using biological materials, so a patient could be exposed to foods that would cause a reaction and potentially prolong proper treatment (Ortolani et al, 1999). Otherwise, risks are limited (Fell, Brostoff, & Pasula, 1988).<br />Case Studies, Testimonials, and Methods of Marketing the Tool<br />During a literature search, few case studies were listed and were not from peer-reviewed sources. The case studies were also quite short in explanation and lacked details needed for scientific review. One case study of a 38-year-old female with severe facial rash, prurutis, fatigue, asthma, and frequent cold, flu, and laryngitis detailed the patient’s health history, physical examination, and diagnostic lab test results, including an ELISA test. Based on the initial ELISA results, the attending physician removed dairy, corn, salmon, apricot, onion, and garlic. All symptoms resolved within 12 weeks, but the rash returned at the 10-month follow-up. The physician decided to use ALCAT to identify any food triggers that would not have been assessed with an IgE/IgG ELISA test; the results showed sensitivities to corn, garlic, dairy, beef, and mustard. The patient was instructed to continue the previous elimination diet but to also eliminate beef, mustard, and other members of the allium family, such as onions and leeks. The rash resolved, and neither it nor any of the other symptoms had returned as of the publication of the case study (Brady, 2010).<br />Several testimonials are available on the ALCAT.com webpage. The testimonials provide examples of how ALCAT has helped patients decrease symptoms related to food sensitivities and intolerances, from obesity to multiple sclerosis. The number of testimonials is quite extensive, which may seem as if this tool is very effective, but it is important to keep in mind legitimacy.<br />Much of the ALCAT marketing occurs directly on the ALCAT webpage and is worldwide, with at least one distributor in a multitude of countries. Practitioners can enroll on the ALCAT webpage to become an ALCAT member and provider of the tool, given that they register with the website. However, anyone can order and purchase the food panels, nutritional support, and even at-home and at-work blood withdrawal from the ALCAT webpage (ALCAT, 2011). The access to such an assessment tool seems overly easy and accessible, decreasing prestige. <br />Literature<br />A study was designed to analyze the reproducibility of ALCAT testing. The patients were referred by a qualified general practitioner or physician and were not known by any of those involved in the study. After consent was obtained, 20mL of blood was withdrawn from 10 subjects and divided into 4 equal volumes of 5mL samples for repeated tests. Inclusion criteria was not detailed, but it was noted that 2 of the 10 had no history of allergy but 8 with one of either migraine, gastroesophageal reflux, irritable bowel syndrome, recurrent sinusitis, allergic rhinitis, asthma, eczema, and tension fatigue syndrome. Exclusion criteria consisted of use of antihistamines, antidepressants, corticosteroids, anticoagulants, and diagnosis of leukemia/neutropenia/lymphocytopenia. 130 different antigens (identified in report) were investigated for a total of 1300 analyses performed. Results showed inaccuracy of 4.0576%, which is a statistical norm for biological systems. The overall reproducibility of ALCAT was 92.0678%, indicating that ALCAT is reliable and reproducible; a p-value was not provided (Neetling & Kachelhoffer, 1998). <br />In another study, 56 children (23 girls, 33 boys), aged 0.5-16.0 (mean 7.2 years) with a history of food hypersensitivities were given a provocation diet then their blood collected in a single-blind trial. Comparison of the ALCAT results to skin prick tests (SPTs) and another diagnostic food allergy tool MAST-CLA (multiple allergen simultaneous test-chemiluminescent assay) showed consistency in over two-thirds, justifying the use of ALCAT for further analysis; a p-value was not specified. Both SPT and MAST-CLA investigate IgE reactions and showed highest consistency of children with sensitivities for nuts, grain, carrot, and soybeans. However, ALCAT results showed that tea, apple, cola, and barley were the most frequent causes of food intolerance in children (Buczyłko et al, 1995). This difference is most likely due to ALCAT’s recognition of other reactivity pathways other than IgE and IgG.<br />ALCAT results were used for an elimination diet for 72 patients (45 children and 27 adults) presenting with food intolerances. The best improvements were achieved in adult patients with arthritis (83%), urticaria (75%), bronchitis (70%), gastroenteritis (70%). Children patients had the worse results of least improvement with hyperactivity (32%), rhinitis (47%), and atopic dermatitis (49%); a p-value was not noted. An IgE-mediated reaction due to external environmental factors other than food is most likely the cause of less improvement in rhinitis and atopic dermatitis. The authors of this study claim effectiveness of ALCAT, as there much improvement in the patients’ symptoms when following an elimination diet based on individualized ALCAT results (Myłek, 1995).<br />A further study investigated ALCAT’s affect on identifying food intolerances that are associated with weight gain. Obesity is associated with inflammation of adipose tissue due to chronic activation of the innate immune system, leading to further weight gain, insulin resistance and diabetes. This inflammation may be caused by food intolerances, so the authors hypothesized that the adherence to a food intolerance elimination diet would improve weight in refractory patients. ALCAT was used to identify the triggers of food intolerance and was used to create the elimination diet. Patients were obese and experienced difficulty losing weight on reduced calorie diet, as well as multiple symptoms such as gastrointestinal reflux, chronic fatigue, headache and other chronic disorders associated with food sensitivities. The trial was for 12 weeks of following ALCAT diet plans. A significant decrease in body weight (from 91.37 ± 10.56 to 74.6 ± 6.76 kg), total body fat percent (from 37.1 ± 7.16 to 27.66 ± 6.52%), and body mass index (32.1 ± 3.8 to 26.1 ± 2.63 kg/m2) was observed; p-values were not provided. Most weight lost was from fat. The authors concluded that ALCAT was reliable in structuring a diet to eliminate symptoms such as obesity, gastrointestinal reflux, chronic fatigue, headache and other chronic disorders associated with food sensitivities (Akmal et al, 2009).<br />A separate study examined the degree of correlation between ALCAT and the Gold Standard oral double blind challenge (DBC). 19 subjects were recruited with symptoms of IBS, atopic eczema, allergic rhinitis, and/or migraine headaches. Their blood samples were tested against 50 food extracts at the beginning and at the end of the 8-week trial. Each participant was to complete a diary card rating the severity of symptoms on a 0 to 4 scale. An elimination diet consisting of 3 ALCAT positive and 3 ALCAT negative foods were eliminated in the first 2 weeks, followed by reintroduction of 1 of the 6 eliminated foods each subsequent week for 6 weeks. An increase in symptoms >40% was considered significant. Of the ALCAT positive foods, 79.3% matched with DBC; ALCAT negative foods matched 83.4% with DBC; no p-value noted. These findings suggest that ALCAT is reliable in identifying trigger foods for individuals (Fell et al, 1988).<br />Another study, by Sandberg and Pasula, consisted of a control group (A) and an experimental group (B), with 25 subjects in each group. The inclusion criteria of group A was asymptomatic, member of the University of Miami Hurricanes football team with a nutritionally balanced diet, no history of food sensitivities, and within the ages of 18-25; group B inclusion criteria was symptomatic with history of food sensitivities and within the ages of 18-25. Group A was all males, whereas group B consisted of 18 males and 7 females. Of 225 reactions, 5 positive from group A (mean 6.56%) and 48 positive from group B (mean 10.80%); p-values were not indicated. The percentage of reactions was significantly higher for group B; therefore, ALCAT can discriminate between negative and patient populations (Sandberg & Pasula, 1988).<br />Additional studies have also found results supporting the safety, efficacy, and reproducibility of ALCAT. Cabo-Soler at the University of Valencia reported that DEXA studies determined more adipose tissue loss than muscle loss in weight loss subjects following isocaloric food elimination diets based on ALCAT results. Another study at the University of Cape Town showed that ALCAT had reproducibility of 94.94%, and a Norwegian study reported ALCAT to be >90% reproducible (Akmal et al, 2009).<br />Despite such positive results of previous ALCAT studies, others state that ALCAT, like many other allergy testing tools, are of obscure theoretical basis and the methods lack technical and clinical validation (Kleine-Tebbe & Herold, 2010). One review stated that “the ALCAT Test is a sophisticated version of the previous “Leukocytotoxic testing”, which was stopped in the United States after the American Academy of Allergy, Asthma and Immunology (AAAAI) statements” (Wüthrich, 2005). The AAAAI proclaimed that there is currently no scientific proof of the efficacy of cytotoxicity test in diagnosing food allergy, and that several controlled studies indicate that the test is ineffective (Ortolani et al, 1999).<br />Several studies showing negativity towards ALCAT are referenced in articles and reviews, but they rarely detail the methods and results of the studies. One such study was performed in the Allergy Unit at Groote Schuur Hospital in 1994 by the University of Cape Town Respiratory Unit, the University of Cape Town Gastrointestinal Unit, and the Allergy Clinic at the Red Cross Children’s Hospital in patients with asthma, eczema and irritable bowel syndrome. The Allergy Society of South Africa stated that “the ALCAT’s predictive value was found to be extremely poor and not of benefit in identifying the trigger of the patient’s symptoms.” The patients, who were followed up by a doctor and by a dietician, did not show any improvement while using the diets based on ALCAT results (Allergy Society of South Africa, 2009).<br />Furthermore, testing a vast list of test agents without obtaining the subject’s history of possible food triggers or even exposure to foods in the fixed panels is neither economical nor useful for diagnosis. As a result, advice and treatments may be improper, the use is not evidence-based, and the patients are not helped. In some patients, such as those with an inhalant allergy, misinterpretation of ALCAT results may lead to inappropriate and unnecessary dietary restrictions (Allergy Society of South Africa, 2009).<br />Each of these studies did not follow the most rigorous of trial designs: a double blind randomized control trial. Randomized control trials oftentimes are of small sample sizes, but a small sample size suggests more susceptibility to the effects of unmeasured variables and imprecise estimates of group difference (Haas, Aickin, & Vavrek, 2010). A study population too small prohibits further analysis beyond the observation of trends; therefore, a population size of at least 50 participants would suffice (Weber Hellstenius, 2009). Bias is introduced when expectation influences outcomes in patients aware of their treatment; therefore the patients should be blinded as to whether their diets are based on their ALCAT results or are in the control group (Haas et al, 2009)<br />In addition, these studies failed to provide p-values of their published results. Further studies should include statistics for scientific reputability. The studies also did not mention the cost effectiveness, as ALCAT testing ranges from $425 through $1099, not including additional support or kits (Haldeman & Dagenais, 2010). The length of the study needs to be at least 8 weeks for a proper collection of baseline blood samples, the elimination diet, and follow-up blood samples (Fell et al, 1988). The studies should then be published in peer-reviewed journals for to maintain standards and for credibility (Wüthrich, 2005).<br />Appropriate Patient Selection Criteria<br />Since ALCAT has not been found to be unsafe or cause serious side effects or illness, this tool could be used on any individual. The symptoms that the ALCAT webpage claims to help ease through elimination diets based on the ALCAT results could be applied to just about everyone (ALCAT, 2011). Previous studies have selected children and adults, symptomatic and healthy, obese and healthy weight, and men and women populations. Patients that should be excluded from ALCAT testing would include those on blood thinners, such as heparin and warfarin, and those with blood-clotting disorders, such as hemophilia and hypercoagulability; these conditions may compromise the patient’s health during or after blood withdrawal (Neetling & Kachelhoffer, 1998). Patients with blood-borne pathogens should notify any practitioners and healthcare professionals of their current health status, so that the blood can be handled safely and appropriately without endangering the health of others (Pasula, 1993).<br />Conclusion<br />The ALCAT company website references clinical studies, editorial articles, and case studies, but mainly provides only the abstracts of articles in non-peer-reviewed journals. Databases such as PubMed only list few papers, which are difficult to access and most do not even provide the abstract (Wüthrich, 2005). Hence, more studies are needed to investigate ALCAT as a scientifically supported food allergy-testing tool.<br />Positive testimonials, case studies, and literature suggest the potential advantage of ALCAT over ELISA, skin prick tests, and other IgE and IgG tests. However, the efficacy of a tool may be overestimated since many trials with negative outcomes are not published. More controlled studies, particularly double blind randomized control trials with large subject populations and statistics, need to be performed to further explore ALCAT, its efficacy, and its safety. In the meantime, healthcare practitioners should be mindful that ALCAT might not be successful in all patients, potentially delaying appropriate treatment (Ortolani et al, 1999).<br />References<br />Akmal M, Khan SA, & Khan AQ. (2009). HYPERLINK quot;
http://www.alcat.com/assets/File/The%20Effect%20of%20The%20ALCAT%20Test%20Diet%20Therapy%20for%20Food%20Sensitivity%20in%20Patients%20With%20Obesity%20-%20Middle%20East%20Journal%20of%20Family%20Medicine%20-%20April%202009,%20Volume%207,%20Issue%203.pdfquot;
quot;
_blankquot;
quot;
The Effect of The ALCAT Test Diet Therapy for Food Sensitivity in Patient’s With Obesityquot;
The Effect of The ALCAT Test Diet Therapy for Food Sensitivity in Patient’s With Obesity. Middle East Journal of Family Medicine, 7(3).<br />ALCAT. (2011). The Technology. Retrieved from http://www.alcat.com/thetechnology.html<br />Allergy Society of South Africa. (2009). Position Statement on ALCAT and IgG Allergy & Intolerance Tests. Retrieved from http://www.allergysa.org/pdfs/intolerance_tests.pdf. <br />Brady DM. (2010). Case Study: Food Intolerance. Clinical Rounds in Functional and Nutritional Medicine.<br />Buczyłko K, Obarzanowski T, Rosiak K, Staśkiewicz G, Fiszer A, Chmielewski S, & Kowalczyk J. (1995). Prevalence of food allergy and intolerance in children based on MAST CLA and ALCAT tests. Rocz Akad Med Bialymst, 40(3):452-6.<br />Fell PI, Brostoff J, & Pasula MI. (12-16 November 1988). HYPERLINK quot;
http://www.alcat.com/assets/File/HIGH%20CORRELATION%20OF%20THE%20ALCAT%20TEST%20RESULTS%20WITH%20DOUBLE%20BLIND%20CHA.pdfquot;
quot;
_blankquot;
High Correlation of the ALCAT Test Results with Double-Blind Challenge (DBC) in Food Sensitivity. Annals of Allergy. 5th Annual Congress of the American of Allergy and Immunology, Los Angeles, CA.<br />Haas M, Aickin M, & Vavrek D. (2009). A preliminary path analysis of expectancy and patient-provider encounter in an open-label randomized controlled trial of spinal manipulation for cervicogenic heachache. Journal of Manipulative and Physiological Therapeutics, 33(1), 5-13.<br />Haldeman S, & Dagenais S. (2010). Choosing a treatment for cervicogenic headache: when? what? how much?. The Spine Journal, 10, 169-171.<br />Kleine-Tebbe J, & Herold DA. (2010). Inappropriate test methods in allergy. Hautarzt, 61(11):961-6. German. <br />Myłek D. (1995). ALCAT Test results in the treatment of respiratory and gastrointestinal symptoms, arthritis, skin and central nervous system. Rocz Akad Med Bialymst, 40(3):625-9.<br />Neetling WML, & Kachelhoffer AM. (1998). Reproducibility of the antigen leucocyte cellular antibody test (ALCAT). Retrieved from http://www.alcat.com/assets/File/REPRODUCIBILITY%20OF%20THE%20ANTIGEN%20LEUCOCYTE%20CELLULAR%20ANTIBODY%20TEST.pdf.<br />Ortolani C, Bruijnzeel-koomen C, Bengtsson U, Bindslev-jensen C, Björkstén B, Høst A, Ispano M, Jarish R, Madsen C, Nekam K, Paganelli R, Poulsen L, & Wüthrich B. (1999). Controversial aspects of adverse reactions to food. Allergy, 54(1): 27–44. doi:10.1034/j.1398-9995.1999.00913.x.<br />Pasula MJ. (1993). The ALCAT test: in vitro procedure for determining food sensitivities. Folia Med Cracov, 34(1-4):153-7.<br />Sandberg DH, & Pasula MJ. (12-16 November 1988). HYPERLINK quot;
http://alcat.com/Images/Pdf/A_Comparison_Of_The_Alcat_Test_For_Food_Reactions.pdfquot;
quot;
_blankquot;
A comparison of the ALCAT test for food reactions amongst 2 population sub-groups. Annals of Allergy. 5th Annual Congress of the American of Allergy and Immunology, Los Angeles, CA.<br />Weber Hellstenius, S.A. (2009). Recurrent neck pain and headaches in preadolescents associated with mechanical dysfunction of the cervical spine: a cross-sectional observational study with 131 students. Journal of Manipulative and Physiological Therapeutics, 32(8), 625-634.<br />Wüthrich B. (2005). Unproven techniques in allergy diagnosis. J Investig Allergol Clin Immunol, 15(2): 86–90.<br />