Cheese in a sack is a specific Croatian indigenous semi hard cheese. The main characteristic of this cheese is ripening and keeping in sheep's clothing (Mišina), which gives it a typical and very appreciated, spicy flavor and aroma. Cheeses obtained by similar technology and ripening are also produced in Turkey (Tulum cheese), Lebanon (darfiyeh), Bosnia and Herzegovina (sack cheese), Montenegro (sack cheese) and eastern Algeria (bouhezza cheese). In Croatia, cheese in a sack has been produced traditionally for many years, without the use of commercial starter cultures, which can result in differences in taste, consistency and microbiological quality.Therefore, there is a growing trend of finding and selecting of new natural indigenous potential starter cultures from traditional products, which will be adapted to the growth in certain specific traditional products, and will have such metabolic capabilities that will significantly affect the quality and safety of products. The hypothesis is that the cheese in a sack with the addition of probiotic bacteria is different from the control cheese in a sack without the addition of probiotic cultures. In literature, there is no data available on the addition of probiotic bacteria in a sheep cheese in a sack. The addition of probiotic bacteria in cheese in a sack presumed differences are expected in the following: microbiological safety of cheese in a sack, the intensity of proteolysis during the ripening of cheese in a sack, the chemical profile of aromatic volatile compounds and fatty acids, sensory evaluation of cheese in a sack and shortene dripening of cheese in a sack. To confirm this hypothesis, the following research objectives were chosen: to determine the effect of probiotic strains Lactobacillus plantarum B and Lactococcus lactissubsp. Lactis S1 isolated from cheese in a sack on microbiological, chemical profile of volatile compounds and fatty acids during ripening, chemical properties, proteolytic changes and sensory evaluation of cheese in a sack in various stages of ripening (15, 30,45 days). The survey was conducted in the area of Knin on the family farm that produces cheese in a sack in the traditional way. For the production of cheese, whole raw sheep milk of the breeds of sheep called Dalmatian pramenka was used. With the addition of probiotic strains Lactobacillus plantarum B and Lactococcus lactis S1 and cheese with mixed probiotic culture (B + S1) and the control cheese without the addition of culture were manufactured. Cheese ripening lasted for 45 days. To explore cheeses in a sack during ripening, samples were collected on zero (gruš), 15, 30 and 45 days of ripening.Before each opening, the sack was placed in a specially designed airtight container in which the air is replaced with an inert nitrogen gas (anaerobic conditions). In such conditions cheese samples were taken by sterile gloves through the "door opening". After sampling and re-closing, the sack was returned to normal conditions of ripening. A total of 4 cheese in a sack were monitored on microbiological, chemical, proteolytic changes and chemical profile of volatile compounds and fatty acids during ripening, and sensory properties of the finished product. The chemical parameters were determined by standard methods. The possible occurrence of pathogenic bacteria Staphylococcus aureus, Listeria monocytogenes, bacteria of the genus Salmonella and yeasts and molds was determined by standard microbiological methods. In order to confirm the presence of applied probiotic strains Lactobacillus plantarum B and Lactococcus lactis S1 in the manufactured samples of probiotic cheese in a sack, apart from their identification on selective plates, the biochemical identification by API and MALDI-TOF method was carried out. For thee valuation in the primary proteolytic changes of cheese, polyacrylamide gel electrophoresis (urea-PAGE) was used. Proteolysis index was calculated by determining the densitometric intensity bands of particular casein fractions and their ratios. The stage of secondary proteolytic changes was found by determination of the water soluble nitrogen (WSN) in relation to the total nitrogen (TN). Chemical analysis of the profile of volatile compounds in cheese in a sack, was conducted by gas chromatography, mass spectrometry (GC-MS), including the use of HS-SPME method for the isolation. Analysis of fatty acid composition of cheese in a sack is determined by gas chromatography with a flame ionization detection (GC-FID). The prepared methyl esters of fatty acids were analyzed according to HRN ISO 5508: 1995 standard, using the gas chromatograph 7890B (Agilent Technologies, Lake Forest, USA), with a capillary column HP88 length of 100 m, a diameter of 0.25 mm and thicknes stationary phase 0.22 micrometers (AgilentTechnologies, Lake Forest, USA), split-splitless injector (temperature 250 C, split ratio1:50) and flame ionization detector (temperature 280 C). Sensory evaluation of cheese was performed by the method of Internal Rules for the evaluation of milk and milk products of the Institute for Dairy, at Faculty of Agriculture, University of Zagreb, where the scoring system with a maximum score of 20 was used. All the statistical analyses have been conducted by using the software package SPSS version 17.0.0 (SPSS Inc., Chicago, IL, USA). The data were statistically processed by using the Tukey's honest significance test and the dominant population of microorganisms during specific phases of production was determined. The results of microbiological, chemical and sensory analysis of cheeses from in a sack indicate that the number of live probiotic bacteria Lactobacillus plantarum B and Lactococcus lactis subsp. Lactis S1 used in the production of probiotic cheeses survived during technological process of cheese production in a great number and that number and that number remained the same during 45 days of ripening. The applied production technology favored the survival of probiotic cultures. The addition of probiotic bacteria has positively influenced the chemical composition of the produced cheese (moisture content, fat and protein), profile of volatile compounds and microbiological validity and sensory properties of cheese in a sack. From the results obtained, it can be concluded that the cheese in a sack is suitable matrix for selected probiotic strains, in particular the application of probiotic cultures are Lactobacillus plantarum B and Lactococcus lactissubsp. Lactis S1 in the production of cheese in a sack the authenticity of the product was kept, and the improvement of qualitative and functional properties of cheese in a sack was included.