Cow's Milk Allergy

Allergen Data Collection - Update: Cow's Milk (Bos domesticus)
Internet Symposium on Food Allergens 4(1): 19-106 (2002) [http://www.food-allergens.de]

4 Diagnostic Features of Cow's Milk Allergy

[Family History / Maternal Factors] [Humoral Parameters] [Cellular Parameters]
[Gastrointestinal Parameters] [Test Significance] [Other Features]

Family History / Maternal Factors References

Family History

Subjects / Follow-up Manifestation of CMA Family History
of Atopic Disease Ref.

children (siblings with CMA) in 33% positive (1)

formula fed infants (5th day to 3 months) in 40%* positive (2)

formula fed infants (5th day to 3 months) in 13%* negative (2)

29 children with severe CMA (1 to 10 months) in 89% (1 parent)
in 50% (both parents) (3)

91 children (8 months) with gastrointestinal symptoms in 34% (14%) (4)

57 children (8 months) with extraintestinal symptoms in 53% (5%) (4)

12 infants (birth to 5 years) persistent** (a) in 83% (5)

26 infants (birth to 5 years) resolved within 1-2 years (b) in 38% (5)

26 children (3 years) persistent in 80% (6)

60 children (3 years) tolerance acquired in 20% (6)

*significance P <0.001
**symptoms at onset predominantly gastrointestinal, at the end of the study increased frequency of wheezing, constipation, and delayed reactions
(a, b) multiple food intolerance in a) 92%, and b) 12%, respectively

(3) occurence of severe CMA in a pair of identical twins and HLA-identical siblings

(4) family history of CMA in paranthesis

(6) 86 consecutive children with IgE-mediated or non-IgE-mediated CMA (median age at diagnosis 4 months)

(1) Gerrard et al. 1973
(2) Vandenplas & Sacre 1986
(3) Schwartz et al. 1987
(4) Ventura & Greco 1988
(5) Iacono et al. 1998c
(6) Carroccio et al. 2000b

Maternal Parameters in Breast Milk

Mothers from IgG IgA TGF-beta-1 HLA-DR # Total No. of
Leukocytes TNF-alpha IFN-gamma Ref.

6 infants with CMA (-)* (1)

65 infants with IgE-mediated CMA (-)* (3)

37 with non-IgE mediated CMA (+)* (3)

36 infants with CMA (-)*** (+) (2)

24 healthy infants (+)*** (-) (2)

48 infants with challenge-proven CMA (-)** (4)

27 infants with challenge-proven CMA (0.25-8.0 months) (-) (+/-) (5)

*in colostrum, ** in colostrum and breast milk, ***significance p=0.012
(-) lower, (+) higher values, and (+/-) no difference as compared to controls
# expression on breast milk macrophages

(2) asymptomatic mothers

(3) TGF-beta-1 positive correlation to beta-LG spec. IgA and CAS spec. IgG, negative correlation to SPT and lymphocyte stimulation with beta-LG or CAS

(4) total and cow's milk specific IgA; levels of specific IgA positively correlated with levels of total IgA but not with the development of CMA in the infants

(5) spontaneous and mitogen-induced TNF-alpha and IFN-gamma production of human milk leucocytes

(1) Savilahti et al. 1991
(2) Järvinen et al. 1999a
(3) Saarinen et al. 1999b
(4) Järvinen et al. 2000a
(5) Järvinen et al. 2000b

Maternal Serum IgG
(1) Mothers of infants who a) developed allergy or b) presented no symptoms:
Statistically lower serum IgG anti- beta-LG levels in a) than in b) (P <0.001) (1) Casimir et al.1989

Humoral Parameters References

Risk Factors for Specific Serum IgE
Significant risk factors for the presence of cow's milk specific IgE in 75 infants with IgE-mediated reactions to cow's milk (mean age of 6.7 months)*:
- long breast-feeding
- exposure to cow's milk at the maternity hospital
- breast-feeding during the first 2 months at home either exclusively or combined with infrequent exposure to small amounts of cow's milk (1) Saarinen & Savilahti 2000
* see Footnote (1)

Specific Serum IgE
Positivity and mean values of cow's milk specific serum IgE:

Patients / Reference (1) (2) (3)

with history of CMA (+) in 71%

cow's milk tolerant children (+) in 27%

cow's milk DBPCFC positive 34 kU/L* 3.9 kU/L**

cow's milk DBPCFC negative 1.7 kU/L* 0.6 kU/L**

(+) increased IgE levels, *significance P<0.0001, **significance P<0.001

(1) 69 children with food intolerance, IgG levels seemed to parallel IgE levels, no differences in IgA levels in allergic and control subjects

(2) 196 children and adolescents with atopic dermatitis (90% family history of atopic diseases)

(3) 107 children with atopic dermatitis

(1) Dannaeus et al. 1977
(2) Sampson & Ho 1997
(3) Niggemann et al. 1999b

Specific IgE, Persistent Allergy
Significantly elevated levels of milk- and CAS- specific IgE in children with persitent CMA (age of >9 years) as compared to children with CMA at the age of <3 years (RAST) (1) (1) Sicherer & Sampson 1999

Total / Specific IgE, SPT, and Persistent Allergy
26 children with persistent CMA had significantly elevated levels of total serum IgE, and cow's milk and cow's milk proteins specific IgE, and higher frequency of positive SPT to cow's milk and proteins (86 consecutive children with IgE-mediated or non-IgE-mediated CMA, median age at diagnosis 4 months) Carroccio et al. 2000b

Native / Denatured CAS, Specific IgE, Persistent Allergy
36 children with CMA: a) 11 became clinically tolerant, and b) 25 had persistent CMA; (6 controls without CMA). Compared to group b) children of group a) had significantly higher ratios of specific IgE antibodies against linear (denatured by reducing agent) than against conformational (native) epitopes of alpha-S1 and beta-CAS; none of group a) children developing tolerance and 9 of group b) children with persistent CMA possessed IgE
to linearized beta-LG (immunodot-blots) Vila et al. 2001

CAS Specific B Cell Epitopes, Persistent Allergy

2 IgE-binding regions of alpha-S1-CAS (aa 69-78 and aa 173-194) detected by 67% and 100% of sera from patients with persistent CMA (>9 years of age) but by none of children less than 3 years of age who are likely to outgrow CMA. No differences in IgG binding observed (24 children with CMA) (1)

6/10 children with persistent CMA detected a peptide from alpha-S1-CAS (aa 69-78) while none of 10 children who outgrew CMA had IgE binding to this epitope (immunodot-blot) (3)

3 IgE binding regions on beta-CAS and 6 on kappa-casein detected by the majority of 15 children with CMA (age of 4-18 years) with high levels of cow's milk specific IgE, but not by sera from children with CMA (age of <3 years) with low levels of specific serum IgE, who are likely to outgrow CMA (2)

(1) Chatchatee et al. 2001a
(2) Chatchatee et al. 2001b
(3) Vila et al. 2001

alpha-LA Specific B Cell Epitopes, Persistent Allergy
4 IgE-binding regions of alpha-LA detected with sera from 11 patients with persistent CMA (4-18 years of age, IgE to cow's milk >100 kU(A)/L) none of these detected by sera from 8 children <3 years of age (IgE to cow's milk <30 kU(A)/L) who are likely to outgrow CMA (SPOTs membrane technique) (1) (1) J ärvinen et al. 2 001

beta-LG Specific B Cell Epitopes, Persistent Allergy
7 IgE-binding regions of beta-LG detected with sera from 11 patients with persistent CMA (4-18 years of age, IgE to cow's milk >100 kU(A)/L) only 3 of these regions detected by sera from 8 children <3 years of age (IgE to cow's milk <30 kU(A)/L) who are likely to outgrow CMA (SPOTs membrane technique) (1) (1) J ärvinen et al. 2 001

Specific IgE, Immediate Reactors, Tolerance
69 IgE- sensitized immediate reacting children with CMA (median age of 24 months) median study period of 2 years: 22% developed clinical tolerance, had lower specific IgE levels at the beginning and the end of study period, and significant fall in SPT reactivity (1) (1) Hill et al. 1993b

beta-LG Specific B Cell Epitopes, Immediate and Delayed Type CMA
8 immediate type patients with CMA (systemic reactions) and 6 delayed type patients with CMA (skin reactions) recognized same B cell epitope (beta-LG aa 95-113), no difference in IgE- binding peptide pattern (RAST inhibition, Pin-ELISA) (1) (1) Heinzmann et al. 1999

Specific Serum IgG and IgA
Percentage of positivity of specific serum IgA and IgG antibodies:

Specificity IgA IgG Ref.

alpha-LA a) 43% b) 44% a) 57% b) 69% (1)

beta-LG a) 71% b) 50% a) 43% b) 75% (1)

beta-LG a > b (2)

beta-LG a, c > b (3)

BSA a > b (2)

BSA a, c > b (3)

CAS a) 86% b) 44% a) 86% b) 69% (1)

CAS a > b '' (3)

pooled alpha-LA, beta-LG, CAS a) 71% b) 38% a) 57% b) 63% (1)

NS = no significant differences

(1) children (age of 3 months to 6 years): a) 7 with CMA (cutaneous symptoms), b) 16 with CMA (gastrointestinal symptoms)

(2) a) 10 infants fed cow's milk- based formula, b) 10 infants fed a CAS hydrolysate formula until the age of 9 months

(3) 129 children a) cow's milk formula fed, b) CAS hydrolysate formula fed, c) breast fed during the first 3 days of life, otherwise exclusively breast fed, follow-up for 2 years ('' at 8 and 12 months)

(1) Bottaro et al. 1992
(2) Vaarala et al. 1995
(3) Juvonen et al. 1999

Specific IgE and IgG Subclass, IgA, Ratios

spec. Ig Cow's Milk CAS beta-LG alpha-LA

IgE a > b (4)
+ (8)
a) 90%, b) 0% (9) a > b (4)
a) 50%, b) 0% (9)
a) 20%, b) 0% (9)

IgE/IgG a > b (4) a > b (4)

IgE + IgG a) 80% (9) a) 40% (9) a) 20% (9)

IgG1 a > b (4); + (8) c > d, e, f** (6); + (8)

IgG4 NS (1)
a > b > c (2) a > b > c (2)
a > b (4)
a) 80%, b) 30% (9) a > c; b > c (2)
+ (8)
a) 80%, b) 20% (9) a) 10%, b) 10% (9)

IgE/IgG1 a > b (4) a > b (4)

IgE/IgG4 a > b (4) a > b (4)

IgG (+) (3)
(-) NS (5) NS (4)
a) 90%, b) 50% (9)
NS (4)
NS (7)
a) 70%, b) 40% (9) a) 60%, b) 10% (9)

IgE + IgG + IgG4 70% (9) 40% (9) 0% (9)

IgA (-) (3)

(+) increase, (-) decrease, (NS) no significant differences
**ratios of IgG1/IgG, IgG1/IgG3 and IgG1/IgG4 same tendency

(1) no relation to provocation test in 14 children with immediate reactions to cow's milk, 15 cow's milk tolerant children

(2) children with immediate type CMA, SPT positive to a) cow's milk (n=20), b) cow's milk and whey hydrolyzed formula (n=17), c) cow's milk, whey and CAS hydrolyzed formulas (n=13)

(3) 21children with challenge proven CMA

(4) a) 18 children with CMA, b) 11 children acquired tolerance

(5) 28 adults with CMA (aged from 16 to 58 years)

(6) c) children with CMA predominantly gastrointestinal , or d) skin symptoms of immediate-onset, e) children with untreated coeliac disease and f) healthy children

(7) a) 12 children with persistent CMA up to age of 5 years, b) 26 controls

(8) 15 adults with CMA (average age of 39.5 years)

(9) positivity in a) 10 children with CMA (age of 7 months to 68 months) and b) 10 age-matched non-allergics

(1) Björkstén et al. 1983
(2) Schwartz 1991
(3) Tainio & Savilahti 1990
(4) James & Sampson 1992
(5) Stoger & Wüthrich 1993
(6) Saalman et al. 1995
(7) Iacono et al. 1998c
(8) Little et al. 1998
(9) Szabó & Eigenmann 2000

Symptoms and Prevalence of Specific IgE
Positivity of cow's milk specific serum IgE in 148 children with CMA according to symptoms:

Symptoms IgE Symptoms IgE

Respiratory 100% Persisting diarrhea 33%

Eczema 71% Severe colics 27%

Urticaria / Anaphylaxis 56% Total 48%

Vomiting 47% Gastrointestianl 33%

Failure to thrive 33% Extraintestinal 72%

Ventura & Greco 1988

Serum Eosinophilic Cationic Protein (ECP)
After 4 weeks of elimination diet; measurement of ECP before oral cow's milk challenge, 27 hours and 1 week after in 28 cow's milk allergic children (age of 5.8 to 43 months): Increased, transient ECP serum levels during challenge in patients with skin manifestations but not in patients with gastrointestinal symptoms (1)
After elimination diet; determination of ECP before milk challenge test, as well as 2 and 24 hours after it in 35 cow's milk allergic children (average age of 16 months; 6-49 months): Basic ECP level significantly higher than in control group; ECP level significantly decreased 2 hours after milk challenge test, after 24 hours back to basic ECP level; no significant differences in ECP levels of children with (n=10) and without clinical reactions (n=25), either before or after challenge test (2) (1) Suomalainen et al. 1994b
(2) Hidvegi et al. 2001

Soluble IL-2 Receptor
Elevated serum levels of soluble IL-2 receptor in 16 children with non- IgE mediated CMA and in 8 children with IgE mediated CMA as compared to 19 children with other IgE-mediated food intolerance (1) (1) Blanco Quiros et al. 1993

IL-12 and sCD30
Increased serum IL-12 levels and normal sCD30 levels in children with CMA while pollen-sensitized children had normal IL-12 and higher sCD30 levels than controls; no differences in patients with asthma or allergic dermatitis (11 children with CMA, open elimination-challenge test, SPT, RAST) (1) Blanco Quiros et al. 1999

Specific TABM
Elevated serum levels of T-cell derived antigen- binding molecules (TABM) specific for alpha-LA, beta-LG, and CAS in 6 to 7 of 15 adults with CMA (1) (1) Little et al. 1998

Eosinophil-related Markers
In a girl (at age of 12 months to 3 years) with CMA and multiple food allergies: high total eosinophil count, increased eosinophil activity, low IFN-gamma : IL-5 ratio, poor wheight gain, increasing respiratory symptoms (1) (1) Nilsson et al. 1999

Cellular Parameters References

Lymphocyte Subclasses, Antigen Expression

Patients T-Cells B-Cells PBMC not specified Ref.

29 children with severe CMA (NS) HLA-A, -B, -DR (1)

7 children with CMA and atopic dermatitis (+)* CLA+ (2)

children with CMA (+) CD8+ (3)

37 children with CMA (+)* HLA-DQ7 (4)

24 children with CMA (0.4-10 months) (-)* CD8+ (+)* total No.
(+)* CD19+ (5)

9 children with IgE-CMA (+)* alpha4beta7 integrin (NS) CD3+CD4+
(-)* CD3+CD8+ (6)

15 fed with cow's milk formula (+)* PCNA (7)

7 breast fed children (+)* CD23+ (7)

12 patients with and without CMA CD4+ (8)

6 patients with CMA (+) CD1d (9)

(+) increase, (-) decrease, * significant, (NS) no significant difference

(1) preparations from unstimulated PBMC

(2) in vitro stimulation with CAS

(3) stimulation with alpha s1-CAS

(4) majority of HLA-DQ7 positive patients presented a high humoral response rather than cellular response (stimulation with beta-LG)

(5) challenge proven patients, compared to healthy controls (no stimulation)

(6) mean age of 28 months (7 months to 9.3 years), beta-LG stimulated PBMC

(7) children with IgE- mediated CMA (3-11 months of age), PCNA expression >/=10% as specific and sensitive marker of CMA in cow's milk fed infants, low cow's milk antigen diets are related with reduced lymphocyte reactivity in whey hydrolyzed fed and breast fed infants (stimulation with beta-LG)

(8) patients with atopic dermatitis; stimulation with CAS, alpha-LA, and beta-LG

(9) expression of CD1 in duodenal biopsy: CD1d positive cells found in lamina propria during symptomatic and asymptomatic periods, 6 healthy controls virtually devoid of CD1d expression; localization of CD1d positive cells in areas where B cells, plasma cells and dendritic cells were present; positive correlation between the numbers of CD1d(+) and CD19(+) cells in the lamina propria

CLA - cutaneous lymphocyte antigen (responsible for skin homing)
PCNA - proliferating cell nuclear antigen (1) Schwartz et al. 1987
(2) Abernathy-Carver et al. 1995
(3) Nakajima et al. 1996
(4) Camponeschi et al. 1997
(5) Jarvinen et al. 1998
(6) Eigenmann et al. 1999
(7) Papadopoulos et al. 1999
(8) Schade et al. 2000
(9) Ulanova et al. 2000

T-Cells, Cell Surface Markers
Cow's milk protein specific T-cell clones (TCCs) were established from blood of infants with a) CMA and atopic dermatitis, from b) atopic controls (atopic dermatitis, without CMA), from c) nonatopic controls, and d) from infants with CMA after spontaneously developed tolerance:

Cell-Surface Marker CD25 CD30 CD26

Expression Levels a > b, c, d a > b, c, d a < c, d

(1) Schade et al. 2002

Lymphocyte* / PBMC** Proliferation

Patients / Stimulation with Cow's Milk
Proteins beta-LG BSA CAS Ref.

17 children with CMA (+) (+) (1)*

children with CMA (challenge proven) (+) (2)*

a) children with CMA
b) children with CMA (immediate type, RAST positive) a) (+)
b) (-) a) (+)
b) (-) (3)**

children with CMA and atopic dermatitis (+) (4)**

a) children with CMA (gastrointestinal symptoms)
b) children with CMA (skin or no symptoms) a > b (5)*

10 children with CMA NS'' NS (6)**

a) <5 years, b) >6 years of age a > b (7)**

a) 10 infants fed cow's milk- based formula
b) 10 infants fed a CAS hydrolysate formula a > b a > b a > b'' (8)**

a) 27 children with IgE mediated CMA
b) 9 children with milk induced enterocolitis syndrome a) (+)
a vs b: NS (9)*

a) 22 patients with cow's milk responsive atopic eczema
b) 66 patients with atopic eczema (non-responsive) a > b (10)**

(+) higher stimulation index or proliferation, (NS) no significant differences

(1) significant proliferation with at least one milk antigen in 15 patients

(2) in children without specific IgE

(3) 3 children with CMA and tension- fatigue sydrome (cow's milk RAST scores in a) negative or slightly positive)

(4) as compared to children with immediate allergic symptoms and controls

(5) 44 children with CMA (mean age of 16 months) after 2-4 weeks of elimination diet, proliferation response abrogated after clinical challenge

(6) as compared to control group ('' stimulation with whey hydrolyzed formula and proteins), lower stimulation with hydrolyzed formula

(7) 22 children with CMA and atopic dermatitis, proliferative response decreased rapidly after elimination diet

(8) fed until the age of 9 months ('' stimulation with alpha-CAS)

(9) a) as compared to control group (significant, but extensive overlapp), group a) also responded to soybean antigen

(10) age of 16-67 years (median 28 years)

(1) Endre & Osvath 1975
(2) Tainio & Savilahti 1990
(3) Kondo et al. 1992
(4) Kondo et al. 1993
(5) Suomalainen et al. 1994a
(6) Eigenmann et al. 1995
(7) Iida et al. 1995
(8) Vaarala et al. 1995
(9) Hoffman et al. 1997
(10) Werfel et al. 1997b

Lymphocyte Transformation
Lymphocyte transformation test a) before and b) 30 days after elimination of cow's milk from the diet: a) significantly increased lymphoblastogenesis (P <0.01), b) no differences in 19 children with CMA (1) (1) Brarda et al. 1989

CBMC Proliferation, IFN-gamma
Stimulation of cord blood mononuclear cells (CBMC) with cow's milk proteins: pronounced proliferation of cells stimulated with alpha-LA, beta-LG, and alpha-CAS; preferentially reduced IFN-gamma levels in individuals with positive parental allergic history (39 randomly selected newborns) (1) (1) Szepfalusi et al. 1997

Cytokine Production by Lymphocytes

Patients / Cytokines IFN-gamma TNF-alpha IL-4 IL-5 IL-10 IL-13 Ref.

a) immediate- reacting
b) late- reacting
c) milk tolerant a < c, b* (1)

a) children with CMA
b) children who acquiered tolerance a < b < c (2)

children with CMA (+) (3, 4)

a) children with CMA
b) children who acquiered tolerance a > b (4)

a) immediate- reacting
b) late- reacting a) (+)
b) (-) (5)

children with atopic dermatitis (milk responsive) (-) (6)

a) children with CMA (cutaneous symptoms)
b) children with CMA (predominantly digestive symptoms)
d) children who acquiered tolerance a > b > c, d ('') (7)

31 children with CMA (-)* (-)* (8)

a) 6 infants with CMA
b) 6 infants without CMA a < b a > b a > b a > b (9)

a) 22 immediate-reacting children
b) 29 late-reacting children
d) cow's milk tolerant children a, b > d a > b (10)

* significant, c) healthy control group, (+) positive response

(1) 75 (a) and 17 (b) children with CMA and 59 (c) tolerant children (age of 1-9 years) (stimulation with beta-LG)

(2) 22 children

(3, 4) stimulation of PBMC with cow's milk proteins

(5) lower thresholds of stimulation in a) as compared to b)

(5) 50 cow's milk allergic children (age of 2-60 months) (DBPCFC positive) with atopic dermatitis, after DBPCFC difference in IFN-gamma generation abolished

(6) IL-4 production of CD4+ CAS specific T-cell clones (compared to house dust mite sensitive patients)

(7) 83 children, measured in whole blood cultured with cow's milk proteins, day 1 ('') followed by TNF-alpha degradation, day 5: secretion peak in group b)

(8) challenge proven children with either skin or gastrointestinal symptoms or both compared to healthy controls (age of 0.12-11.2 months), unstimulated PBMC and mitogen- induced production

(9) infants with atopic dermatitis (age of 3.8-12.3 months); cow's milk protein-specific TCC derived from PBMCs stimulated with CAS, alpha-LA, and beta-LG; 2 patients with TCC reactivity to CAS and whey proteins and 2 patients with exclusive reactivity to CAS in both groups; CMA in infants with atopic dermatitis associated with production of TH-2 cytokines by circulating antigen-specific CD4+ T cells; significant correlation with IL-4 of both IL-5 and IL-13 production

(10) stimulation of PBMC with cow's milk; IL-10 concentrations increased in response to DBPCFC in challenge-positive children

(1) Hill et al. 1993a
(2) Suomalainen et al. 1993a
(3) Heyman et al. 1994
(4) Benlounes et al. 1996
(5) Sutas et al. 1997
(6) Werfel et al. 1997b
(7) Benlounes et al. 1999
(8) Österlund et al. 1999
(9) Schade et al. 2000
(10) Sütas et al. 2000

Cytokine Secreting Cells in Blood and Duodenal Mucosa
Frequency of spontaneously cytokine secreting mononuclear cells in the

blood duodenal mucosa

INF-gamma a, b > c a > c

IL-4 b > a > c a > c

IL-5 a, b > c a = c

IL-10 a, b > c a < c

Children with a) CMSE, b) CMA, and c) age matched controls
Cytokine secreting cells more frequently in duodenal mucosa than in the blood (1) Hauer et al. 1997

PBMC, Migration Inhibition Factor
In vitro assay of lymphocyte migration inhibition factor (MIF), stimulation of peripheral blood lymphocytes with beta-LG: significant higher MIF production in 24 children with CMA than in control subjects; most of 18 children recovered from CMA had negativ assay (1) (1) Ashkenazi et al. 1980

Lymphocytes, Suppressor Activity
Decreased suppressor activity of isolated lymphocytes induced by either Concanavalin A or cow's milk in 10 children with CMA as compared to controls and patients who acquired cow's milk tolerance (1) (1) Suomalainen et al. 1993b

Cell Mediated Cytotoxicity
Antibody- dependent cell- mediated cytotoxicity (ADCC) to beta-LG- coated cells rather induced in most sera of children with CMA and predominantly gastrointestinal symptoms than in sera of children with skin reactions (immediate- type), children with untreated coeliac disease, or healthy children; ADCC reactivity of individual sera correlated with their IgG1 antibody levels (1) (1) Saalman et al. 1995

Homing Receptor Expression
Stimulation of a) cord blood mononuclear cells and b) peripheral blood mononuclear cells (at age of >3 months) with alpha-S1 CAS:
a) higher percentage of alpha-E-beta-7-positive T cells than in healthy controls; no difference in cord blood T-cell proliferation; CLA was not induced on T cells
b) no induction of alpha-E-beta-7-positive T cells; CLA was expressed on T cells
(4 infants with atopic dermatitis and CMA) (1) Kohno et a. 2001

Gastrointestinal Parameters References

Salivary IgA
158 healthy mature infants at birth: Salviary anti-CAS IgA was significantly higher (P <0.05) in high risk infants than in no risk or low risk infants; salviary anti-CAS IgA values correlated with maternal allergy, but not with paternal allergy (1) (1) Renz et al. 1990

Pancreatic Enzymes
children with CMA (median age 3 months) fed with a) a hydrolyzed CAS- based formula or b) a soy- protein based formula: No significant difference in pancreatic secretion between both groups for any of the enzymes studied (trypsin, chymotrypsin, lipase, and phospholipase) during diet of 6 weeks (1) (1) Carroccio et al. 1997

Duodenal Fluid, Specific IgE and IgD
increased levels of cow's milk protein (and soybean agglutinin) specific IgE and IgD in basal and pancreozymin- stimulated duodenal fluid in 13 children with various intestinal diseases (1) (1) Freier et al. 1983

Jejunal Fluid, Hyaluronic acid, Albumin
Jejunal fluid levels of hyaluronan (hyaluronic acid) and albumin increased after milk perfusion challenges in 5 adults with CMA (DBPCFC positive, SPT and RAST negative, lactose tolerant) as compared with control group (1) (1) Bengtsson et al. 1996

Small Intestine Mucosa, IgE and IgM Plasma Cells
local reaginic reaction after ingesting cow's milk: increased mucosal IgE and IgM plasma- cells, increased degranulation of mast cells, staining of connective tissue and basement membranes with antisera to IgG and C3 complement in 2 cow's milk sensitive infants (1) (1) Shiner et al. 1975

Small Intestine Mucosa and Serum, Alkaline Phosphatase
Levels of alkaline phosphatase (ALP) after cow's milk protein challenge: Significant depletion in upper jejunal mucosa tissue and serum in infants with clinical and histological reactions (n=10); tissue ALP depressed in 3/5 patients with histological but no clinical reactions to cow's milk (1) (1) Iyngkaran et al. 1995

Small Intestinal IgE Plasma Cells, Specific Serum IgE

Patients / Cow's Milk Specific IgE Plasma Cells Serum IgE Ref.

16 children with CMA (+) in 56% (+) in 38% (1)

15 without CMA (+) in 6.7% (+) in 13% (1)

(1) elimination / challenge proven CMA

(1) Schrander et al. 1993a

Intestinal Total Immunoglobuline Secreting Cells
Intestinal immune responses after diagnostic milk provocation:

Patients / No. of Secreting Cells IgM IgA IgG Ref.

a) with CMA (acute urticaria) (+) (-) (-) (1)

b) with CMA (gastrointestinal symptoms) (+) (+) (-) (1)

c) with CMA (skin and gastrointestinal symptoms) (+) (+) (+) (1)

d) 13 with persistent CMA (+) (+) (+) (2)

e) 24 acquired tolerance (-) (-) (-) (2)

d) 27 with CMA (age of 9-69 months) (+) (+) (+) (3)

(+) significant increase during challenge, (-) no increase

(1) IgM and IgA responses: in group b) > a)

(3) increase in all isotypes associated with clinically positive cow's milk challenge; specific antibody secreting cells against beta-LG and CAS (and gliadin) increased in IgM class only

(1) Isolauri et al. 1990
(2) Isolauri et al. 1992
(3) Suomalainen et al. 1992

Intestinal Eosinophils, Lymphocytes, Mast Cells

Patients Eosinophils Lymphocytes TIA-1** Mast Cells Ref.

12 children with CMA (+) in 58% (-) (1)

47 children with coeliac disease (+) in 60% (-) (1)

children with CMA and chronic diarrhea (+)* (2)

21 children with CMA/CMI (+) in 38% (+)* (3)

35 children with gluten intolerance (+) in 27% (+)* (3)

10 children with CMA/CMI (+)* (+)* (4)

* significant, (+) increase, (-) decrease
**TIA-1 (= cytotoxic granule-associated protein) expressing lymphocytes

(1) in lamina propria of jejunum

(2) in lamina propria of duodenal mucosa

(3) cellular infiltration of small intestinal mucosa

(4) number of TIA1- expressing intraepithelial lymphocytes (IEL) and the TIA1/IEL ratio in patients on cow's milk-free diet of various duration, negative correlation between the TIA1/IEL ratio and the duration of the diet (duodenal biopsies)

(1) Kosnai et al. 1984
(2) Challacombe et al. 1986
(3) Kaczmarski et al. 1989
(4) Hankard et al. 1997

Intestinal ECP, MBP, Histamine, VCAM-1

Patients ECP* MBP* Histamine* VCAM-1** Ref.

5 adults with CMA (+) (+) (1)

14 patients with cow's milk-sensitive enteropathy (+) (+) (2)

(+) increased, (-) decreased, *intestinal secretion, **expression on mononuclear cells

(1) DBPCFC positive, SPT and RAST negative, lactose tolerant patients (perfusion challenges with milk, CAS, and whey)

(2) Challenge positive, SPT and RAST negative patients, endoscopic duodenal biopsy

ECP = eosinophil cationic protein

MBP = eosinophil major basic protein

VCAM-1 = vascular cell adhesion molecule-1

(1) Bengtsson et al. 1997
(2) Chung et al. 1999

Intestinal Epithelial Cells, CD23 Expression
CD23 expression on intestinal epithelial cells increased in 3 children with CMPI (age < 1 year) associated by high levels of specific IgE Kaiserlian et al. 1995

Fecal alpha-1 Antitrypsin, TNF-alpha, ECP, IgE
Indicators of intestinal inflammation in jejunal fluid after cow's milk challenge:

Patients alpha-1 Antitrypsin TNF-alpha ECP IgE Ref.

13 children with CMA (gastrointestinal symptoms) (+)* (-) (-) (1)

a) positive DBPCFC with cow's milk
b) negative DBPCFC with cow's milk (+) in 43%
(+) in 11% (+)**
(-) (+)***
(-) (2)

15 children with CMA (+) in 58% (3)

(+) significant increase after challenge, (-) no increase
*in challenge positive children
**particularly in delayed type patients, ***particularly in immediate reactors

(2) children with atopic eczema

(3) out of 26 atopic infants with confirmed food allergy; all 9 patients with increased fecal concentration of alpha-1-antitrypsin had positive challenge with cow's milk while only 6 in those with normal alpha-1-antitrypsin concentration (fecal samples collected before elimination diet and 3 months later)

(1) Kapel et al. 1999
(2) Majamaa et al. 1996
(3) Majamaa et al. 2001

Gastrointestinal Permeability

Urinay Recovery / Test Substances Permeability Alteration
after cow's milk challenge Ref.

polyethylenglycol (PEG) * (1)

lactulose/mannitol excretion ratios (+) (3)

cellobiose/mannitol excretion ratios (+) (4)

lactitol/mannitol excretion ratios a > b (5)

Jejunal Biopsy / Test Substances

horseradish peroxidase (HRP) (+)* (2)

*significantly changed, (+) increased, (-) decreased

(1) 16 children with CMA (immediate- type), greatest alteration in children with most severe symptoms

(2) 15 children with CMA (age of 1-24 months), jejunal transepithelial fluxes

(3) 51 children with CMA (skin symptoms and patients with gastrointestinal symptoms), 3 days after challenge

(4) 32 children with CMA (age of 3-84 months), 24 hours after challenge

(5) children with symptoms suggestive of CMA (age of 0.5-168 months): a) 95 children with proven CMA and b) 105 controls (challenge negative); defining a cut-off value intestinal permeability exhibited a 68% sensitivity and a 77% NPV for CMA;. highest sensitivity (70%) at ages 6-12 months; abnormal intestinal permeability in 80% of CMA children with digestive manifestations, in 43% with extra-digestive, 68% with mixed and 40% with anaphylactic manifestations; lactitol/mannitol ratio correlated negatively with age in control group, no correlation in CMA group

(1) Falth-Magnusson et al. 1986
(2) Heyman et al. 1988
(3) Jalonen 1991
(4) Troncone et al. 1994
(5) Kalach et al. 2001

Protein / Allergen Absorption
Concentrations in blood serum samples

Patients human alpha-LA bovine beta-LG Ref.

17 children with CMA (age of 3-78 months) 0.3 to 2 µg/L (in 29%) (1)

20 infants (followed up to 8 months) 3-4 days after birth 31 µg/L
at 1 month 6 µg/L
at 2 months 2 µg/L
at >3 months trace amounts after weaning
1 week 7 µg/L (in 38%)
2 weeks 4 µg/L (in 21%) (2)

(1) 24 h after cow's milk challenge

(2) median serum levels (per g alpha-LA or beta-LG given per kg body weight)

(1) Husby et al. 1990
(2) Kuitunen et al. 1994

Diagnostic Significance of Tests References

SPT, Atopy Patch Test (APT), RAST

Patients / Reference (1) (2) (3) (4) (5) (6) (7) (8)

Age (months) adults 2-36 2-24 50* 62* <24 21*

a) acute onset 14 36 22

b) delayed onset 18 50

SPT - positive
(cutoff point, 3 mm) a) 67%
b) (-) a) (+) 55% 14%

sensitivity
specificity
PPV
NPV 66%
100%
100%
28% 96%
51%
66%
93%

APT - positive a) (-)
b) 89% b) (+) 44%

sensitivity
specificity
PPV
NPV

RAST - positive
(cutoff point, >0.35 kU/L) 26%

sensitivity
specificity
PPV
NPV 79%
80% 75-100%
71%
60-67%
83-100% 100%
30%
57%
100% 85%
38%
61%
71%

* mean age, (-) tendency of negative results, (+) association to positive results

(1) 14 children with immediate reactions to cow's milk, 15 cow's milk tolerant children

(2) 21 adults with cow's milk / egg allergy (DBPCFC, 5 different RAST systems)

(3) 183 children with CMA and atopic dermatitis (DBPCFC or open challenge, 54% challenge positive)

(4) 54 children with CMA and atopic dermatitis

(5) 430 food allergic children and adolescents (labial food challenge positive) age from 0.2 to 20 years

(6) 54 of 109 DBPCFC positive to cow's milk (study population: 196 food allergic children and adolescents with atopic dermatitis, age from 0.6 to 17.9 years)

(7) 72 children with CMA (challenge proven)

(8) 107 children with atopic dermatitis (47 DBPCFC positive) age from 5 months to 12 years

(1) Björkstén et al. 1983
(2) Norgaard et al. 1995
(3) Isolauri & Turjanmaa 1996
(4) Kekki et al. 1997
(5) Rance et al. 1997
(6) Sampson & Ho 1997
(7) Majamaa et al. 1999
(8) Niggemann et al. 1999b

SPT, Atopy Patch Test (APT), RAST (continued)

Patients / Reference (9) (10) (11) (12) (13)

Age (months) 2-11 1-192 <12 13* 6.3-7.5

a) acute onset 100 125 75 58

b) delayed onset 76 17 60

SPT - positive
(cutoff point, 3 mm) a) (+)

sensitivity
specificity
PPV
NPV 69%
91%
79%
85%
79% 99%
38%
56%
97% 78%
69%
81%
64% 61%
76%
71%
67%

APT - positive a) (+)

sensitivity
specificity
PPV
NPV 18%
87%
40%
69% 47%
96%
95%
51% 37%
77%
61%
56%

RAST - positive
(cutoff point, >0.35 kU/L) a) (+)

sensitivity
specificity
PPV
NPV 58%
88%
70%
81% 85%
56%
61%
83% 84%
38%
70%
59% 72%
49%
58%
64%

Serum ECP - positive
(cutoff point, >15 µg/L)

sensitivity
specificity
PPV
NPV 27%
74%
67%
34%

* mean age, (-) tendency of negative results, (+) association to positive results

(9) 301 children with suspected CMA (176 DBPCFC positive)

(10) 310 children with suspectd CMA (43% challenge positive), specifity of SPT in children less than 2 years of age was 91%

(11) 170 children with suspected CMA age <1 year (mean 4.8 months); 4-year follow-up; SPT and RAST: at least one positive result for whole milk and/or major milk proteins (alpha-LA, beta-LG, or CAS)

(12) 45 of 71 DBPCFC positive to cow's milk (study population: 98 children with atopic dermatitis and suspected food allergy, age from 2 months to 11.2 years); combination of positive APT with evidence of specific IgE or with a positive SPT resulted in a PPV of 100%

(13) 239 children with suspected CMA (mean age of 6.9 months); 49% challenge positive; with a cut-off level in SPT of 8 mm sensitivity and NPV decreased (to 19% and 55%, respectively) while specificity and PPV increased (to 98% and 92%, respectively); using protein fractions (beta-LG, bovine serum albumin, CAS) in APT sensitivity and NPV were 26% and 57%, respectively, while specificity and PPV increased (to 92% and 77%, respectively); with a cut-off level in RAST of >3.5 kU/L sensitivity and NPV decreased (to 25% and 57%, respectively) while specificity and PPV increased (to 98% and 94%, respectively); serum ECP measured after challenge (day 4); with a cut-off level of 24.7 µg/L serum ECP sensitivity decreased to 13% while specificity, PPV, and NPV increased (to 98%, 93%, and 37%, respectively)

(9) Vanto et al. 1999
(10) Sporik et al. 2000
(11) Garcia-Ara et al. 2001
(12) Roehr et al. 2001
(13) Saarinen et al. 2001

Predictive Decision Points of Specific IgE
It should be noted that pedictive decision points are significantly affected by specific conditions within each study population (e.g. atopic dermatitis or selection criteria)

Patients / Reference (1) (1) (1) (1) (2) (2) (3) (3)

Age (months) 62* 62* 62* 62* <12 <12 13* 13*

a) acute onset 75 75 100% -

b) delayed onset - 100%

Predictive Decision Point (kU(A)/L)** <0.8 <1.0 >23 >32 >2.5 >5 >17.5 >17.5

sensitivity
specificity
PPV
NPV 98%
41%
95%
95%
48%
90%
58%
94%
90% 51%
98%
95% 48%
95%
90%
69% 30%
99%
95%
64% 22%
96%
86%
54 17%
96%
75%
63%

Patients / Reference (4) (4) (5) (5)

Age (months) 6.9* 6.9* 45.6* 45.6*

a) acute onset 58 58

b) delayed onset 60 60

Predictive Decision Point (kU(A)/L)** >0.7 >3.5 >15 >32

sensitivity
specificity
PPV
NPV 45%
87%
78%
61% 25%
98%
94%
57% 57%
94%
95%
53% 34%
100%
100%
44%

* mean age, ** CAP system FEIA

(1) 54 of 109 DBPCFC positive to cow's milk (study population: 196 food allergic children and adolescents with atopic dermatitis, age from 0.6 to 17.9 years)

(2) 170 children with symptoms suggesting immediate-type CMA (age <1 year, mean 4.8 months); 4-year follow-up

(3) 45 of 71 DBPCFC positive to cow's milk (study population: 98 children with atopic dermatitis and suspected food allergy, age from 2 months to 11.2 years); combination of positive APT with evidence of specific IgE in a PPV of 100%

(4) 239 children with suspected CMA (age of 6.3-7.5 months); 49% challenge positive

(5) 100 children and adolescents with suspected IgE-mediated food allergy (age of 3 months to 14 years); 61% with atopic dermatitis, appr. 50% with asthma, and 90% with atopic family history, 21/62 DBPCFC positive to cow's milk

(1) Sampson & Ho 1997
(2) Garcia-Ara et al. 2001
(3) Roehr et al. 2001
(4) Saarinen et al. 2001
(5) Sampson 2001

Predictive Decision Points of SPT
It should be noted that pedictive decision points are significantly affected by specific conditions within each study population (e.g. atopic dermatitis or selection criteria)

Patients / Reference (1) (1) (2) (2)

Age (months) <24 31* 6.9* 6.9*

a) acute onset 37% 58 58

b) delayed onset 5% 60 60

Predictive Decision Point (wheal diameter in mm) >6 >8 >6 >8

sensitivity
specificity
PPV
NPV
100%
100% 37%
93%
83%
60% 19%
98%
92%
55%

* mean age

(1) 310 children with suspected CMA (age of 1-192 months; 120 children < 24 months); 55% of challenges were positive, 37% negative, and 8% inconclusive (open oral food challenges)

(2) 239 children with suspected CMA (age of 6.3-7.5 months); 49% challenge positive; SPT with cow's milk formula

(1) Sporik et al. 2000
(2) Saarinen et al. 2001

Cut-Off Levels: Specific IgE vs. SPT
In EAST 62 children from 640 children with suspected food allergy (<2 years of age) were CMA positive according to specific IgE levels grade 3+ and 4+ (>3.5 AEU/mL) while 63 children had IgE-mediated CMA by SPT (100% diagnostic SPT level: >6 mm) but not by EAST;
in CAP RAST 4 children from 127 children with suspected food allergy (<2 years of age) were CMA positive according to specific IgE levels grade 4, 5, and 6 (>17.5 kU(A)/L) while 11 children had IgE-mediated CMA by SPT (100% diagnostic SPT level: >6 mm) but not by CAP RAST Hill et al. 2001

SPT, IgE and Oral Challenge
Diagnostic tests in comparison with oral challenge test in 11 children with CMA (1-15 years of age):

SPT RAST MAST Case History

Sensitivity 85% 71% 71%

Specificity 100% 100%

Match 60% 81% 81% 63%

Roger et al. 1994

SPT, IgE, APT, ECP and Oral Challenge
Positivity of tests in 239 children challenged with cow's milk at a mean age of 6.9 months:

Oral Challenge SPT RAST APT ECP

118 positive 61% 45% 26% 21%

121 negative 24% 13% 8% 13%

Combination of the 4 tests correctly classified 73% of the infants with a sensitivity of 76% and a specificity of 67% (cut-off levels: SPT >/ = 3 mm; RAST cow's milk-specific IgE >/ = 0.7 kU/L; ECP >/ = 20 µg/L) Saarinen et al. 2001

Clinical History, IgE, Oral Challenge
Children with atopic dermatitis:

Diagnosis of CMA by (1) (2)

History only - 18%

RAST only 0% 0%

History and RAST 57% 27%

Oral challenge 43% 73%

No. of patients 7 11

(1) Eigenmann et al. 1998
(2) Eigenmann & Calza 2000

SPT and DBPCFC
Significant differences in SPT (wheal sizes) between cow's milk allergic or tolerant individuals (DBPCFC) (P < 0.001); SPT cut-off values mean diameter 5 mm / surface area of wheal 29 mm² Eigenmann & Sampson 1998

Skin Tests, RAST, Histamine Release and Lymphocyte Stimulation
Positive results with cow's milk (and alpha-CAS*)

Test a) b)

SPT 57% 0%

Patch Test* 33% 0%

RAST 59% 33%

Histamine Release 55% 17%

Lymphocyte Proliferation 77% 17%

a) 22 children with CMA (positive challenge test)
b) 12 non- milk- allergic controls with atopic dermatitis
Panel of tests detected 21/22 children positive and 5/12 false- positive (1) Rasanen et al. 1992

Specific Serum IgG

Patients / Reference (1) (2) (3)

Age 16-58, median 26 years 1-48 (1-72) months

cow's milk specific IgG

NS NS

beta-LG specific IgG

sensitivity
specificity 89%
85%

NS no diagnostic significance

(1) 28 adults with CMA

(2) 218 healthy children, 205 with CMA, 96 with other (atopic) diseases (commercial betalactotest)

(3) 702 infants divided into six groups of different feeding (breast fed, infant formula fed)

the shorter the breast feeding period and the earlier cow's milk formula is introduced, the higher the IgG levels

(1) Stoger & Wüthrich 1993
(2) Iacono et al. 1995b
(3) Keller et al. 1996

SPT and RAST
41 children with suspected CMA (age of 3 months to 13 years; mean 2.6 years): 32% SPT positiv, 61% IgE positiv; concordance of SPT and IgE results in 51% (1) (1) Campbell et al. 1987

Histamine Release, SPT and RAST
26 children with suspected CMA: 77% positive in oral challenge test; patients with urticaria: high degree of correlation between histamine test, RAST and skin test; patients with gastrointestinal symptoms only a few positive results in histamine test, RAST and skin test (1) (1) Prahl et al. 1988

Positivity of Open Challenge and DBPCFC
in children with history of CMA

Patients Open Challenge DBPCFC Ref.

265 children suspected for CMA
(mean age 3 months) 56% (n=155) 44% (n=110) (2)

a) 16 probable immediate reactors (mean age 37 months) a) 62.5% with adverse reactions (up to 2 h after milk exposure) (1)

b) 53 probable delayed reactors (mean age 17 months) b) 28.8% with predominantly gastrointestinal symptoms (2h to 6 days after milk exposure) (1)

(1) Baehler et al. 1996
(2) Kaila & Isolauri 1997

Other Features

Parameters / Subjects Outcome References

Gender of Adults with CMA
34 patients with CMA (aged from 16 to 58 years) Gender: 91% females, 39% of them experienced first symptoms during or soon after a pregnancy;
47% of patients were nonatopic and showed a monovalent sensitization to cow milk proteins Stoger & Wüthrich 1993

Intrauterine Sensitization
A newborn presenting symptoms suggestive of CMA First hour of life: hemorrhagic meconium
Next few days: bloody diarrhea
Day 14: elevated total IgE, specific IgE to cow's milk and an eosinophilia in peripheral blood; symptoms disappeared when milk feed was changed to extensively hydrolyzed casein-formula. Day 30 and at 7 months of age: 2 challenges with cow's milk formula followed by recurrence of vomiting, watery diarrhea and failure to thrive.
Age of 17 months: cow's milk well tolerated well Feiterna-Sperling et al. 1997

Intrauterine Sensitization
An infant with non-IgE mediated CMA Symptoms of food-induced enterocolitis occurred before any oral intake of antigen Kalayci et al. 2000

Sensitization / First Symptoms
118 infants with challenge proven adverse reactions to cow's milk (mean age of 6.7 months) 50 infants showed first adverse symptoms during exclusive breast-feeding (37 had cow's milk specific IgE), and 32 infants were sensitized during exclusive breast-feeding (23 had cow's milk specific IgE) Saarinen & Savilahti 2000

Cow's Milk Exposure
25 children with CMA (age <1 year) Exposure to cow's milk formulas (significantly more often than in control group, p < 0.01):
16 during their first week of life
6 before fifth week of life
3 infants not exposed Stintzing & Zetterstrom 1979

SPT, IgE in Immediate Type CMA
26 (1), 50 (2, 3), and 21 (4) children with IgE mediated acute reactions of CMA Differentiation of 3 groups by positive SPT to:
A) cow's milk only
B) cow's milk and whey hydrolysate formula
C) cow's milk, whey and CAS hydrolysate formula (1, 2, 3)
Significant differences in cow's milk specific serum IgE: A < B < C (1, 2)
Significant differences in beta-LG and CAS specific serum IgE: A < C and B < C (1, 2, 3)
Most significant difference in intensity scores of IgE- binding to CAS and beta-LG in SDS-PAGE immunoblot: A < C (4) (1) Schwartz et al. 1989
(2) Schwartz 1991
(3) Schwartz et al. 1991
(4) Amonette et al. 1993

SPT, IgE and Delayed Reactions
49 infants with clinical history suggestive of CMA, positive SPT and RAST (alpha-LA, beta-LG, and CAS) (age of <6 months):
94% had immediate reactions
6% had delayed type reactions
(>2 h after challenge) Reexamination of 24 children with negative SPT and RAST at 1 year of age after exclusion diet:
79% challenge negative
21% challenge positve (all presenting late reactions at mean of 7 days after milk ingestion) Plaza Martin et al. 2001

Frequencies of IgE- and non-IgE CMA
52 of 101 DBPCFC positive to cow's milk (study population of 139 children with atopic dermatitis suspected for food allergy)

IgE-mediated CMA
(SPT and/or RAST positive) in 88%

non-IgE-mediated CMA
(SPT and RAST negative) in 12%

Niggemann et al. 2001b

Adverse vs Allergic Reactions
9 children with "unequivocal symptoms attributable to cow's milk" CMA in 1 patient, abnormal disaccharide absorption in 3 patients (gastrointestinal and immunoallergic investigations) Davidson et al. 1976

"Residual" Allergic Disease
Reexamination of
a) 53 10-year-old children who manifested CMA before 1 year of age and achieved small-dose tolerance
b) age-matched control group of 90 school children

a) b)

reported milk-related gastro-intestinal symptoms 45% 10%

response of intestinal symptoms* 3/6 7/10

growth retarded "normal"

specific IgA and IgG lower higher

lactose malabsorption 14% 3%

* 4-week blind elimination-challenge test with 1 week of low-lactose milk flour Kokkonen et al. 2001b

SPT: Commercial Extract vs. Fresh Food
4-month-old infant with severe atopic dermatitis and positive skin reaction upon contact with cow's milk Reexamination 1 year later:
SPT with commercial CAS, alpha-LA, and beta-LG solutions all negative; in drop test with pasteurized cow's milk a severe generalized urticaria resulted within a few minutes Di Berardino et al. 2001

5 Therapy of Cow's Milk Allergy
6 Composition of Cow's Milk
7 Allergens of Cow's Milk
8 Isolation & Preparation