10 Stability of Cow's Milk Allergens
 

Treatment	Effects	References
Heat boiling of skimmed milk, 10 min	Reduction of IgE binding to alpha-LA about 50%, to CAS fractions >66% and to beta-LG, BSA and bovine Ig binding abolished (CRIE score)	Gjesing et al. 1986
Pasteurization, Homogenization, Hydrolysis 1) raw untreated, 2) pasteurized or, 3) homogenized and pasteurized cow's milk, and 4) a commercial hypoallergenic hydrolysed CAS infant formula	Similar positive reactions in SPT and DBPCFC to 1), 2) and 3), no reactions to 4) in 5 cow's milk allergic children (immediate- type); tendency of lower thresholds of processed milk	Host & Samuelsson 1988
Heat boiling of milk, 2 and 10 min	Boiling of milk for 10 but not for 2 min eliminated SPT reactivity to BSA and beta-LG, whereas CAS was heat stabile (8 DBPCFC positive adults with CMA)	Norgaard et al. 1996
Heat boiling of milk, 5 min	No difference in IgE- binding to raw and cooked milk from sera of 2 patients with CMA (immuno-dot-blotting)	Werfel et al. 1997a
Heat boiling of extensively hydrolyzed CAS formula (Nutramigen)	Positive SPT to boiled and unboiled formula in a girl with CMA	Nilsson et al. 1999
Heat, Reduction, Hydrolysis a) heat 96°C, pH 8.0 b) carboxymethylation c) limited proteolysis	Thermoaggregated (a) and reduced forms (b) exhibited similar anaphylactic effect on sensitized guinea pigs; allergenic properties of beta-LG appeared unaltered in food hydrolyzates after thermal treatment and limited proteolysis	Gmoshinskii et al. 1990
gamma Irradition alpha-CAS, beta-LG	ELISA inhibition using patients' IgE and rabbit IgGs specific for alpha-CAS and beta-LG, respectively: reduced IgE-binding to irradiated proteins with different slopes of inhibition curves; rabbit IgG-binding increased up to a certain dose of irradiation and then decreased; main bands disappeared in SDS-PAGE  with increasing doses while turbidity increased indicating a decrease of solubility (20 patients with CMA)	Lee et al. 2001
Pepsin Hydrolysis, Pancreatic Enzymes, Whey Proteins hydrolysis with pepsin (pH 2, 3, and 4, for 90 min) followed by hydrolysis with a mixture of pancreatic enzymes (pH 7.5 for 150 min) of beta-LG, alpha-LA, BSA, and bovine IgG	Percent inhibition of IgE-binding from sera of patients with IgE- mediated CMA to native proteins by respective hydrolysates of homologue protein: pepsin hydrolysis at pH 2 pH 3 pH 4 beta-LG* 8% 0% 0% alpha-LA 5% 14% 48% BSA 0% 0% 58% bovine IgG 2.5% 54% 91% (RAST inhibition) * beta-LG is barely hydrolysed by pepsin, but susceptible to pancreatic enzymes	Schmidt et al. 1995
Duodenal Digestion in vitro digestion of cow's milk proteins by duodenal fluid (1) and human trypsins and elastases (2)	Rate of hydrolysis: purified proteins > crude cow's milk or formula; rates of alpha-LA, beta-LG, and CAS digestion were 0.03, 0.12, and 16.1 mg/mL duodenal juice/min (same capacity to hydrolyze the milk proteins in infants with CMPI, celiac disease, or unclassified gastrointestinal disorder)	(1) Jakobsson et al. 1982 (2) Jakobsson et al. 1983
Intestinal Digestion intestinal digestion of beta-LG, alpha-LA and CAS (intestinal cells, 48h)	Lower TNF-alpha production by stimulation of PBMC from cow's milk allergic patients with intestinal digested cow's milk proteins as compared to intact proteins	Benlounes et al. 1996
Cell-Envelop Proteinase specific hydrolysis of CAS and ultrafiltration	Elimination of IgE-binding epitopes by degradation of CAS and removal of whey proteins by ultrafiltration	Alting et al. 1998
Pepsin Hydrolysis, beta-LG pepsin hydrolysis of beta-LG	222 cow's milk intolerant / allergic patients: Higher degree of IgG binding to native beta-LG than to pepsin digested LG as compared to controls; almost complete discrimination between patient groups by cross- reactive experiments (ELISA)	Duchateau et al. 1998
Lactic Acid Fermentation lactic acid fermentation of sterilized cow's milk with a mixed culture of meso- and thermophilic bacteria strains	About 99% reduced antigenicity of whey proteins as compared to raw milk (rabbit pAb, ELISA); allergenicity of alpha-LA and beta-LG only slightly attenuated (Skin Tests)	Jedrychowski & Wroblewska 1999

11 Allergen Sources
 

Reported Adverse Reactions	References
Human Milk Cow's milk taken by the mother precipitated allergic symptoms in 61 breast fed infants (elimination/challenge in mother's diet) (1) Proctocolitis triggered by cows' milk protein transferred to 11 infants via the breast milk; resolution of visible rectal bleeding within 72 to 96 hours after elimination of cow's milk from mother's diet (colonoscopy performed in 5 infants revealed benign eosinophilic proctocolitis) (2)	(1) Gerrard & Shenassa 1983a, 1983b (2) Pumberger et al. 2001
"Non-dairy" Foods Allergic reactions in 6 patients with CMA after ingestion of "non-dairy" products: tofu frozen dessert (2 cases), beef hot dog (2 cases), bologna (2 cases), rice frozen dessert, tuna packed in aqueous solution; contents of milk proteins due to contamination by manufacturing facilities (37-2202 µg/mL) or to unlabeled adding of caseinate (136 µg/mL) (1)	(1) Gern et al. 1991
Cake, Cookie, Pastry Near-fatal anaphylactic reactions in 2 children with CMA (age of 9 and 12 years) after ingestion of a cookie, a cake or pastry containing the allergen (1)	(1) Sampson et al. 1992
Chocolate, Candy Occupational asthma and rhinoconjunctivitis in a chocolate candy worker (positive conjunctival and bronchial challenge tests with lactalbumin) (1)	(1) Bernaola et al. 1994
Several Food Products 9 Adverse reactions to unexpected cow's milk allergens in a) Meatballs containing 1.1% CAS (undeclared) b) Hot dog containing 0.04% CAS (contaminated) c) Recombined ham containing 2.6% (undeclared) d) Sausage containing 1.0% CAS (undeclared) e) Sausage containing 0.06% CAS (contaminated) f) Dark chocolate containing 0.8% CAS (contaminated) g) Ice cream (soy based) containing 0.2% CAS (contaminated) h) Lollipop, strawberry / cream containing 0.2% CAS (undeclared) i) Meringue containing 1.1% CAS (undeclared)	Malmheden Yman et al. 1994
Sorbets Frozen desserts manufactured using the same equipment used for producing ice cream may contain milk allergens: Anaphylaxis in a 2-year-old boy after ingestion of a "pareve"- labeled raspberry sorbet (kosher labeling to select dairy-free foods) (1) Anaphylaxis in a 3-year-old boy within 20 min after ingestion of a lemon sorbet (ca. 113 to 170 g) containing trace amounts of milk allergens (whey proteins: 9 µg/mL)  (2)	(1) Jones et al. 1992 (2) Laoprasert et al. 1998
Sausage Severe reactions after ingestion of sausage containing CAS (texturing agent) (1)	(1) Foucard et al. 1997
Cheese Anaphylaxis after ingestion of cheese in a 23-year old woman (1)	(1) Wüthrich & Hofer 1986
Goat's and Sheep's Cheese Several allergic reactions after ingestion of feta (cheese made from sheep's milk) in a 15-year-old boy, after ingestion of sheep's or goat's cheese in a 25-year old patient, both tolerated ingestion of diary procucts from cow's milk  (1) Allergic reactions after eating goat's cheese and after touching of goat's and sheep's cheese in a 2-year-old girl with tolerance to dairy products from cow's milk (2)	(1) Wüthrich & Johansson 1995 (2) Umpierrez et al. 1999
Sheep's Cheese Several anaphylactic reactions after ingestion of food containing "pecorino" cheese made from sheep's milk in a 5-year-old atopic boy unaffected by cow's milk protein allergy (1)	(1) Calvani & Alessandri 1998
Mozarella / Ricotta / Parmesan Cheese Asthma, urticaria and rhinitis in a boy with atopic dermatitis after ingestion of mozarella cheese made from ewe's and cow's milk; several allergic reactions after ingestion of ricotta cheese containing ewe's milk and parmesan cheese made from cow's milk, respectively (1)	(1) Fiocchi et al. 1999
Baby Food A baby food, cereal flour P, containing alpha- lactalbumin (although labeled guaranteed free of cow's milk), caused failure to thrive and diarrhea, vomiting, and coughing fits in a 22- month- old child with cow's milk allergy (1)	(1) Fremont et al. 1996
Casein in Salmon A 30 year-old woman with CMA (without fish sensitivity) experienced anaphylaxis within 1 hour after ingestion of a slice of bread with salmon, prepacked from a supermarket, the salmon meat was restructured by microbial enzyme (transglutaminase) which crosslinks added CAS and meat proteins, assuming a consumption of 10 to 50 g salmon, about 10 to 50 mg CAS were ingested (1)	(1) Koppelman et al. 1999
Casein-containing Product A 25-year-old with asthmatic symptoms which were work-related to the production process of candy making due to inhalation of Hyfoama DS (bronchial challenge, SPT, RAST); Hyfoama DS is a colorless, water-soluble powder produced from CAS treated with calcium hydroxide solution and then dried. It is used as a substitute for egg white in a variety of baked goods. The manufacturer recommends labeling of the final product as containing “hydrolyzed milk protein.”	(1) Bader et al. 2001
Casein in Tryptone Powder Adult-onset sensitization to CAS in a 44-year-old male scientist after occupational exposure to aerosolized tryptone powder (CAS product used in preparation of culture media); subsequent systemic allergic reaction after ingestion of milk containing candy (SPT)	(1) Vaswani et al. 1999
Human Milk 16/17 of cow's milk allergic children showed allergic symptoms during cow's milk challenge through human milk from asymptomatic mothers (age of children 1.8 to 9.4 months)	(1) Jarvinen et al. 1999b
Infant Formulas	see 12.2 Allergenicity / Safety of Infant Formulas
Pharmaceuticals Anaphylaxis in a 12 months-old boy after cutaneous application of a CAS containing ointment to an inflamed diaper area (1) Severe anaphylactic reaction in a 33-year-old woman during artificial insemination due to sperm- processing medium containing BSA (SPT, RAST) (2) Generalized anaphylactic reaction a few hours after tooth extraction in a 54-year-old woman, hemofibrine (a hemostatic sponge made of bovine fibrin) elicited symptoms (Scratch test, RAST) (3) 3 out of 6 patients with positive RAST against CAS experienced allergic symptoms after taking antibiotics containing CAS (4)	(1) Jarmoc & Primack 1987 (2) Wüthrich et al. 1995 (3) Wüthrich et al. 1996 (4) Hasegawa et al. 2000
"Neutralizing" of Food Allergies 68-year-old woman with systemic mastocytosis  was given "neutralizing" injections of milk and wheat and experienced flushing, palpitations, and lightheadedness with syncope (unproven technique of provocation/neutralization) (1)	(1) Teuber & Vogt 1999

 

Allergens in Products	Content / Products	References
Human Milk 38 mothers	Detectable amounts of immunoactive beta-LG (5-33 ng/mL) in 18/38 human milk samples	Jakobsson et al. 1985
Human Milk 25 mothers	Detectable amounts of beta-LG (5-800 ng/mL) in 40%of human milk samples, presence of symptoms in the infant such as diarrhoea, vomiting, colic, exanthema significantly correlated to high levels of beta-LG in the milk	Axelsson et al. 1986
Human Milk 57 breastfeeding mothers	Detectable amounts of bovine beta-LG (up to 6.4 ng/mL) in 45% of samples; persisted up to 3 days after maternal dietary elimination of cow's milk	Machtinger & Moss 1986
Human Milk 9 breastfeeding mothers	Detectable amounts of bovine beta-LG (0.5-45 ng/mL) in 30% of samples	Host et al.1988
Human Milk 36 samples of human breast milk	Detectable amounts of bovine IgG (mean 3.4 ng/ml)	Maeda et al. 1993
Human Milk 55 breastfeeding mothers (cow's milk allergy in 46 infants, oral challenge)	beta-LG in human milk before and after oral cow's milk load (given after 24 hour milk free diet): beta-LG levels >2 ng/mL in 75% of samples, increased levels in 50% after load	Sorva et al. 1994
Fat Substitutes 13 (16) egg and/or cow's milk allergic patients	Allergenicity of microparticulated egg and cow's milk proteins in fat substitues (Simplesse, Beta IL): No alteration of allergenic potencies in SDS-PAGE immunoblot	(1) Sampson & Cooke 1990 (2) Sampson & Cooke 1992
Flavourings / Seasonings dill-pickle seasoning (containing lactose)	Positive skin test reaction to lactose containing seasoning (1 patient with CMA); CAS and whey proteins identified in this seasoning, and whey proteins in pharmaceutical grade lactose	Watson et al. 1995
Lactose "food-quality" lactose	alpha-LA content in "food quality" lactose 1-5 µg/g (RAST inhibition)	Fremont et al. 1996
Natural Rubber Latex Gloves cow's milk CAS	15/30 commonly marketed natural rubber latex glove brands contained cow's milk CAS (rocket immunoelectrophoresis, ELISA inhibition)	Ylitalo et al. 1999

 

Allergen Depleted Products	Method	References
Cow's Milk depletion of beta-LG	Specific affinity matrix: beta-LG coupled to Sepharose 4B used to remove beta-LG from milk, significant retardation in its elution because reversible polymerization with soluble beta-LG; beta-LG depleted milk proposed as hyposensitizing milk formula to be used by lactating mothers	Chiancone & Gattoni 1993

 

Allergen Free Products	Method	References
Human Milk lactating mothers	No detection of beta-LG and bovine CAS in human milk (SDS-PAGE immunoblotting using mAb)	Restani et al. 2000

12 Food Allergen Labeling
 

Food Allergen	Labeling / Regulation Status	References
International Regulations Milk and milk products	Mandatory labeling of prepackaged food / advisary status (1)	(1) Codex Alimentarius Commission 1999
European Regulations Milk and milk products	Labeling appropriate / recommendation (1) Mandatory labeling of prepackaged food / proposal for directive (2)	(1) Bousquet et al. 1998 (2) European Commission 2001

13 Infant Formulas
14 References