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Effects of milk collection and processing methods on origin and integrity of RNA in milk
Author(s) -
Lemay Danielle G,
Hinde Katie,
Hovey Russell C,
Smilowitz Jennifer T,
Hartono Stella R,
Ventimiglia Frank,
Schmidt Kimberli A,
Lee Joyce W.S.,
Korf Ian,
Barry Peter A,
German J Bruce
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.624.2
Subject(s) - rna , macaque , milking , biology , rhesus macaque , microbiology and biotechnology , chemistry , biochemistry , immunology , gene , zoology , paleontology
Given the potential diagnostic utility of RNA in milk, it is important to establish best practices for its collection and processing. Milk was collected from 5 human participants and 6 rhesus macaques at 2 time points and each macaque underwent a mammary biopsy. Degradation of human milk fat RNA was positively correlated with distance from collection site and time stored in TRIzol at −80 C. RNA integrity also varied by sample type: milk fat RNA was significantly more degraded than RNA from cells in milk or mammary tissue. Intriguingly, there were no differences in RNA degradation in macaque milk collected after 10 mins or 4 hours of accumulation, suggesting that milk fat RNA degradation may not occur in the breast. Origin of RNA was determined by applying our automated method to images of milk stained with acridine orange. Human whole milk contained more cytoplasmic crescents and fewer nucleated cells than macaque milk. A repeated milking protocol increased crescent‐derived RNA in some, but not all, macaque milk samples. Using RT‐PCR, expression of a transcript specific to milk‐producing cells, alpha‐lactalbumin, was significantly higher in milk fat compared to mammary tissue. Milk fat RNA may more accurately portray the RNA profile of milk‐producing cells in vivo compared to RNA from mammary tissue; however, this sample type requires adherence to protocols that minimize RNA degradation. Funded by NIH P51RR000169.